Pharmaceutical Research

, Volume 27, Issue 2, pp 285–295

Bioactive Long-Term Release from Biodegradable Microspheres Preserves Implanted ALG-PLO-ALG Microcapsules from In Vivo Response to Purified Alginate

  • Stefano Giovagnoli
  • Paolo Blasi
  • Giovanni Luca
  • Francesca Fallarino
  • Mario Calvitti
  • Francesca Mancuso
  • Maurizio Ricci
  • Giuseppe Basta
  • Ennio Becchetti
  • Carlo Rossi
  • Riccardo Calafiore
Research Paper

Abstract

Purpose

To assess whether prevention of unexpected in vivo adverse inflammatory and immune responses to biohybrid organ grafts for the treatment of Type I Diabetes Mellitus (T1DM) is possible by superoxide dismutase and ketoprofen controlled release.

Methods

Superoxide dismutase and ketoprofen-loaded polyester microspheres were prepared by W/O/W and O/W methods, embodied into purified alginate-poly-L-ornithine-alginate microcapsules and intraperitoneally implanted into CD1 mice. The microspheres were characterized for morphology, size, encapsulation efficiency, enzyme activity and in vitro release. Purified alginate contaminants were assayed, and the obtained microcapsules were investigated for size and morphology before and after implantation over 30 days. Cell pericapsular overgrowth and expression were evaluated by optical microscopy and flow cytometry.

Results

Superoxide dismutase and ketoprofen sustained release reduced cell pericapsular overgrowth in comparison to the control. Superoxide dismutase release allowed preserving the microcapsules over 30 days. Ketoprofen-loaded microspheres showed some effect in the immediate post-grafting period. A higher macrophage and T-cell expression was observed for the control group.

Conclusions

Microspheres containing superoxide dismutase and ketoprofen may represent novel tools to limit or prevent unpredictable adverse in vivo response to alginate, thus contributing to improve cell transplantation success rates in T1DM treatment.

KEY WORDS

alginate microcapsules biodegradable microspheres diabetes post-grafting response superoxide dismutase and ketoprofen 

REFERENCES

  1. 1.
    Nathan DM. The rationale for glucose control in diabetes mellitus. Endocrinol Metabol Clin North Am. 1992;21:221–35.Google Scholar
  2. 2.
    Hemkens G, Grouven U, Bender R, Gunster C, Gutschmidt S, Selke GW, et al. Risk of malignancies in patients with diabetes treated with human insulin or insulin analogues; a cohort study. Dialettologia. 2009;52(9):1732–44.CrossRefGoogle Scholar
  3. 3.
    Shapiro AM, Lakey JR, Ryan EA, Korbutt GS, Toth E, Warnock GL, et al. Islet transplantation in seven patients with type 1 diabetes mellitus using glucocorticoid-free immunosuppressive regimen. N Engl J Med. 2000;343:230–8.CrossRefPubMedGoogle Scholar
  4. 4.
    Shapiro AMJ, Ricordi C, Hering BJ, Auchincloss H, Lindblad R, Robertson RP, et al. N Engl J Med. 2006;355:1318–30.CrossRefPubMedGoogle Scholar
  5. 5.
    Calafiore R, Basta G. Alginate/poly-L-ornithine microcapsules for pancreatic islet cell immunoprotection. In: Kühtreiber WM, Lanza RP, Chick WL, editors. Cell encapsulation, technology and therapeutics. New York: Birkhäuser; 1999. p. 138–50.Google Scholar
  6. 6.
    Calafiore R. Bioartificial pancreas. In: Dumitriu S, editor. Polymeric Biomaterials. New York: Marcel Dekker; 2002. p. 983–1005.Google Scholar
  7. 7.
    De Vos P, van Hoogmoed GC, van Zanten J, Netter S, Strubbe JH, Buster HJ. Long-term biocompatibility, chemistry, and function of microencapsulated pancreatic islets. Biomaterials. 2003;24:305–12.CrossRefPubMedGoogle Scholar
  8. 8.
    Calafiore R. Alginate microcapsules for pancreatic islet cell graft immunoprotection: struggle and progress towards the final cure for type1 diabetes mellitus. Expert Opin Biol Ther. 2003;3:201–5.CrossRefPubMedGoogle Scholar
  9. 9.
    Calafiore R, Basta G, Luca G, Lemmi A, Montanucci MP, Calabrese G, et al. Microencapsulated pancreatic islet allografts into nonimmunosuppressed patients with type 1 diabetes. Diab Care. 2006;29:137–8.CrossRefGoogle Scholar
  10. 10.
    Orive G, Hernandez RM, Rodrıguez Gascon A, Calafiore R, Ming Swi Chang T, de Vos P, et al. History, challenges and perspectives of cell microencapsulation. Trends Biotechnol. 2004;22:87–92.CrossRefPubMedGoogle Scholar
  11. 11.
    Vizzardelli C, Molano RD, Pileggi A, Berney T, Cattan P, Fenjves ES, et al. Neonatal porcine pancreatic cell clusters as a potential source for transplantation in humans: Characterization of proliferation, apoptosis, xenoantigen expression and gene delivery with recombinant AAV. Xenotransplantation. 2002;9:14–24.CrossRefPubMedGoogle Scholar
  12. 12.
    Korbutt GS, Elliott JF, Ao Z, Smith DK, Warnock GL, Rajotte RV. Large scale isolation, growth, and function of porcine neonatal islet cells. J Clin Invest. 1996;97:2119–29.CrossRefPubMedGoogle Scholar
  13. 13.
    Living Cell Technologies Updates Progress with Diabetes Clinical Trial. http://www.lctglobal.com/latest-news.php (accessed 10/27/09).
  14. 14.
    Open-label Investigation of the Safety and Effectiveness of DIABECELL(R) in Patients With Type I Diabetes Mellitus http://www.clinicaltrials.gov/ct2/show/NCT00940173?term=pig+islets&rank=1 (accessed 10/30/09).
  15. 15.
    Zimmermann H, Shirley SG, Zimmermann U. Alginate-based encapsulation of cells: past, present, and future. Curr Diabetes Rep. 2007;7:314–20.CrossRefGoogle Scholar
  16. 16.
    Ricci M, Blasi P, Giovagnoli S, Rossi C, Macchiarulo G, Luca G, et al. Ketoprofen controlled release from composite microcapsules for cell encapsulation: effect on post-transplant acute inflammation. J Control Release. 2005;107:395–407.CrossRefPubMedGoogle Scholar
  17. 17.
    Korbutt GS, Elliott JF, Radiate RV. Cotransplantation of allogeneic islets with allogeneic testicular cell aggregates allows long term graft survival without systemic immunosuppression. Diabetes. 1997;46:317–22.CrossRefPubMedGoogle Scholar
  18. 18.
    Luca G, Basta G, Calafiore R, Rossi C, Giovagnoli S, Esposito E, et al. Multifunctional microcapsules for pancreatic islet cell entrapment: design, preparation and in vitro characterization. Biomaterials. 2003;24:3101–14.CrossRefPubMedGoogle Scholar
  19. 19.
    Luca G, Calvitti M, Basta G, Baroni T, Neri LM, Becchetti E, et al. Mitogenic effects of Brazilian arthropod venom on isolated islet beta cells: in vitro morphologic ultrastructural and functional studies. J Investig Med. 2003;51:79–85.CrossRefPubMedGoogle Scholar
  20. 20.
    Luca G, Calafiore R, Basta G, Ricci M, Calvitti M, Neri L, et al. Improved function of rat islets upon co-microencapsulation with Sertoli’s cells in alginate/poly-l-ornithine. AAPSPharmSciTech. 2001;2: article 15.Google Scholar
  21. 21.
    Giovagnoli S, Luca G, Casaburi I, Blasi P, Macchiarulo G, Ricci M, et al. Long-term delivery of superoxide dismutase and catalase entrapped in poly(lactide-co-glycolide) microspheres: in vitro effects on isolated neonatal porcine pancreatic cell clusters. J Control Release. 2005;107:395–407.CrossRefPubMedGoogle Scholar
  22. 22.
    Sharkawy AA, Klitzman B, Truskey GA, Reichert WM. Engineering the tissue which encapsulates subcutaneous implants. III Effective tissue response times. J Biomed Mater Res. 1998;40:598–605.CrossRefPubMedGoogle Scholar
  23. 23.
    Johansson U, Olsson A, Gabrielsson S, Nilsson B, Korsgren O. Inflammatory mediators expressed in human islets of Langerhans: implications for islet transplantation. Biochem Bioph Res Commun. 2003;308:474–9.CrossRefGoogle Scholar
  24. 24.
    Omer A, Keegan M, Czismadia E, De Vos P, Van Rooijen N, Bonner-Weir S, et al. Macrophage depletion improves survival of porcine neonatal pancreatic cell clusters contained in alginate macrocapsules transplanted into rats. Xenotransplantation. 2003;10:240–51.CrossRefPubMedGoogle Scholar
  25. 25.
    Basta G, Sarchielli P, Luca G, Racanicchi L, Nastruzzi C, Guido L, et al. Optimized parameters for microencapsulation of islet cells: an in vitro study clueing on islet graft immunoprotecion in type 1 diabetes mellitus. Transpl Immunol. 2004;13:289–96.CrossRefPubMedGoogle Scholar
  26. 26.
    de Vos P, Bucko M, Gemeiner P, Navratil M, Svitel J, Faas M, et al. Multiscale requirements for bioencapsulation in medicine and biotechnology. Biomaterials. 2009;30:2559–70.CrossRefPubMedGoogle Scholar
  27. 27.
    Thanos CG, Calafiore R, Basta G, Bintz BE, Bell WJ, Hudak J, et al. Formulating the alginate-polyornithine biocapsule for prolonged stability: Evaluation of composition and manufacturing technique. J Biomed Mater Res. 2007;83A:216–24.CrossRefGoogle Scholar
  28. 28.
    de Vos P, van Hoogmoed CG, de Haan BJ, Busscher HJ. Tissue responses against immunoisolating alginate-PLL capsules in the immediate posttransplant period. J Biomed Mater Res. 2002;62:430–7.CrossRefPubMedGoogle Scholar
  29. 29.
    King A, Sandler S, Andersson A. The effect of host factors and capsule composition on the cellular overgrowth on implanted alginate capsules. J Biomed Mater Res. 2001;57:374–83.CrossRefPubMedGoogle Scholar
  30. 30.
    Babensee E, Anderson JM, McIntire LV, Mikos AG. Host response to tissue engineered devices. Adv Drug Deliv Rev. 1998;33:111–39.CrossRefPubMedGoogle Scholar
  31. 31.
    Gorbet MB, Sefton MV. Endotoxin: the uninvited guest. Biomaterials. 2005;26:6811–7.CrossRefPubMedGoogle Scholar
  32. 32.
    van Schilfgaarde R, de Vos P. Factors influencing the properties and performance of microcapsules for immunoprotection of pancreatic islets. J Mol Med. 1999;77:199–205.CrossRefPubMedGoogle Scholar
  33. 33.
    Vargas F, Vives-Pi M, Somoza N, Armengol P, Alcalde L, Martí M, et al. Endotoxin contamination may be responsible for the unexplained failure of human pancreatic islet transplantation. Transplantation. 1998;65:722–7.CrossRefPubMedGoogle Scholar
  34. 34.
    Dusseault J, Tam SK, Menard M, Polizu S, Jourdan G, Yahia L, et al. Evaluation of alginate purification methods: Effect no polyphenol, endotoxin, and protein contamination. J Biomed Mater Res. 2006;76A:243–51.CrossRefGoogle Scholar
  35. 35.
    Tama SK, Dusseault J, Polizua S, Menard M, Hallé JP, Yahia L. Impact of residual contamination on the biofunctional properties of purified alginates used for cell encapsulation. Biomaterials. 2006;27:1296–305.CrossRefGoogle Scholar
  36. 36.
    de Vos P, Faas MM, Strand B, Calafiore R. Alginate-based microcapsules for immunoisolation of pancreatic islets. Biomaterials. 2006;27:5603–17.CrossRefPubMedGoogle Scholar
  37. 37.
    Giovagnoli S, Blasi P, Ricci M, Rossi C. Biodegradable microspheres as carriers for native superoxide dismutase and catalase delivery. AAPS Pharm Sci Tech. 2004;5(4): article 51Google Scholar
  38. 38.
    Blasi P, Giovagnoli S, Schoubben A, Ricci M, Rossi C, Luca G, et al. Preparation and in vitro and in vivo characterization of composite microcapsules for cell encapsulation. Int J Pharm. 2006;324:27–36.CrossRefPubMedGoogle Scholar
  39. 39.
    Shibuya T, Watanade Y, Nalley KA, Fusco A, Salafsky B. The BCA protein determination system: an analysis of several buffers, incubation temperature and protein standards. Tokyo Ika Daigaku Zasshi. 1989;47(4):677–82.Google Scholar
  40. 40.
    Marklund S, Marklund G. Involvement of the superoxide anion radical in the autoxidation of pyrogallol and a convenient assay for superoxide dismutase. Eur J Biochem. 1974;47:449–74.CrossRefGoogle Scholar
  41. 41.
    Basta G, Calafiore R. A process for the ultrapurification of alginates. 2009. WO/2009/093184.Google Scholar
  42. 42.
    Bradford MM. A rapid and sensitive for the quantitation of microgram quantitites of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–54.CrossRefPubMedGoogle Scholar
  43. 43.
    Fink PC, Lehr L, Urbaschek RM, Kozak J. Limulus amebocyte lysate test for endotoxemia: investigations with a femtogram sensitive spectrophotometric assay. Klin Wochenschr. 1981;59:213–8.CrossRefPubMedGoogle Scholar
  44. 44.
    Active Standard ASTM F2064. Standard guide for characterization and testing of alginates as starting materials intended for use in biomedical and tissue-engineered medical products. 2006. http://www.astm.org/Standards/F2064.htm.
  45. 45.
    Orive G, Tam SK, Pedraz JL, Hallé JP. Biocompatibility of alginate-poly-l-lysine microcapsules for cell therapy. Biomaterials. 2006;27(20):3691–700.CrossRefPubMedGoogle Scholar
  46. 46.
    Blasi P, Schoubben A, Giovagnoli S, Perioli L, Ricci M, Rossi C. Ketoprofen poly(lactide-co-glycolide) physical interaction. AAPSPharmSciTech. 2007;8(2): article 37.Google Scholar
  47. 47.
    Blasi P, D’Souza S, Selmin F, DeLuca PP. Plasticizing effect of water on poly(lactide-co-glycolide). J Control Release. 2005;108(1):1–9.CrossRefPubMedGoogle Scholar
  48. 48.
    Dembczynski R, Jankowski T. Determination of pore diameter and molecular weight cut-off of hydrogel-membrane liquid-core capsule for immunoisolation. J Biomater Sci Polym Ed. 2001;12(9):1051–8.CrossRefPubMedGoogle Scholar
  49. 49.
    Marikovsky M, Ziv V, Nevo N, Harris-Cerruti C, Mahler O. Cu/Zn superoxide dismutase plays important role in immune response. J Immunol. 2003;170:2993–3001.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Stefano Giovagnoli
    • 1
  • Paolo Blasi
    • 1
  • Giovanni Luca
    • 2
  • Francesca Fallarino
    • 3
  • Mario Calvitti
    • 3
  • Francesca Mancuso
    • 3
  • Maurizio Ricci
    • 1
  • Giuseppe Basta
    • 2
  • Ennio Becchetti
    • 3
  • Carlo Rossi
    • 1
  • Riccardo Calafiore
    • 2
  1. 1.Dipartimento di Chimica e Tecnologia del Farmaco, Faculty of PharmacyUniversity of PerugiaPerugiaItaly
  2. 2.Dipartimento di Medicina Interna (Di.M.I.), Sezione di Medicina Interna e Scienze Metaboliche ed Endocrine, School of MedicineUniversity of PerugiaPerugiaItaly
  3. 3.Dipartimento di Medicina Sperimentale e Scienze BiochimicheUniversity of PerugiaPerugiaItaly

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