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Cell and Tissue Banking

, Volume 13, Issue 4, pp 663–671 | Cite as

Ice-free cryopreservation of heart valve allografts: better extracellular matrix preservation in vivo and preclinical results

  • Kelvin G. M. BrockbankEmail author
  • Katja Schenke-Layland
  • Elizabeth D. Greene
  • Zhenzhen Chen
  • Olaf Fritze
  • Martina Schleicher
  • Renate Kaulitz
  • Iris Riemann
  • Falko Fend
  • Johannes M. Albes
  • Ulrich A. Stock
  • Milan Lisy
Original Paper

Abstract

The purpose of this study was evaluation of an ice-free cryopreservation method for heart valves in an allogeneic juvenile pulmonary sheep implant model and comparison with traditionally frozen cryopreserved valves. Hearts of 15 crossbred Whiteface sheep were procured in Minnesota. The valves were processed in South Carolina and the pulmonary valves implanted orthotopically in 12 black faced Heidschnucke sheep in Germany. The ice-free cryopreserved valves were cryopreserved in 12.6 mol/l cryoprotectant (4.65, 4.65, and 3.31 mol/l of dimethylsulfoxide, formamide and 1,2-propanediol) and stored at −80°C. Frozen valves were cryopreserved by controlled slow rate freezing in 1.4 mol/l dimethylsulfoxide and stored in vapor-phase nitrogen. Aortic valve tissues were used to evaluate the impact of preservation without implantation. Multiphoton microscopy revealed reduced but not significantly damaged extracellular matrix before implantation in frozen valves compared with ice-free tissues. Viability assessment revealed significantly less metabolic activity in the ice-free valve leaflets and artery samples compared with frozen tissues (P < 0.05). After 3 and 6 months in vivo valve function was determined by two-dimensional echo-Doppler and at 7 months the valves were explanted. Severe valvular stenosis with right heart failure was observed in recipients of frozen valves, the echo data revealed increased velocity and pressure gradients compared to ice-free valve recipients (P = 0.0403, P = 0.0591). Histo-pathology showed significantly thickened leaflets in the frozen valves (P < 0.05) and infiltrating CD3+ T-cells (P < 0.05) compared with ice-free valve leaflets. Multiphoton microscopy at explant revealed reduced inducible autofluorescence and extracellular matrix damage in the frozen explants and well preserved structures in the ice-free explant leaflets. In conclusion, ice-free cryopreservation of heart valve transplants at −80°C avoids ice formation, tissue-glass cracking and preserves extracellular matrix integrity resulting in minimal inflammation and improved hemodynamics in allogeneic juvenile sheep.

Keywords

Cryopreservation Heart valve Allogeneic Transplantation 

Notes

Acknowledgments

We like to thank Dr. Holly Neaton and Ilka Degenkolbe for their excellent technical assistance. This study was financially supported by the German Research Foundation (Deutsche Forschungsgemeinschaft STO 359/7-1) and Organ Recovery Systems, Inc.

References

  1. Aberle T, Huber AJT, Schleicher M, Brockbank KGM, Stock UA (2011) Impact of different cryopreservation technologies on heart valve leaflet cell viability. In: 15th annual Hilton Head workshop, regenerative medicine: innovations for clinical applications, abstractGoogle Scholar
  2. Bechtel JF, Stierle U, Sievers HH (2008) Fifty-two months mean follow up of decellularized SynerGraft-treated pulmonary valve allografts. J Heart Valve Dis 17:98–104PubMedGoogle Scholar
  3. Brockbank KG, Lightfoot FG, Song YC, Taylor MJ (2000) Interstitial ice formation in cryopreserved homografts: a possible cause of tissue deterioration and calcification in vivo. J Heart Valve Dis 9(2):200–206PubMedGoogle Scholar
  4. Brockbank KGM, Chen Z, Song YC (2010) Vitrification of porcine articular cartilage. Cryobiology 60:217–221PubMedCrossRefGoogle Scholar
  5. Brockbank KGM, Heacox AE, Schenke-Layland K (2011a) Guidance for removal of fetal bovine serum from cryopreserved heart valve processing. Cells Tissues Organs 193:264–273PubMedCrossRefGoogle Scholar
  6. Brockbank KGM, Wright GJ, Yao H, Greene ED, Chen ZZ, Schenke-Layland K (2011b) Allogeneic heart valve preservation—allogeneic heart valve storage above the glass transition at −80°C. Ann Thorac Surg 91:1829–1835Google Scholar
  7. Campbell LH, Brockbank KGM (2010) Cryopreservation of porcine aortic heart valve leaflet-derived myofibroblasts. Bioprocess Biobanking 8(4):211–217CrossRefGoogle Scholar
  8. Joudinaud TM et al (2008) Redo aortic root surgery for failure of an aortic homograft is a mayor technical challenge. Eur J Cardiothorac Surg 33:989–994PubMedCrossRefGoogle Scholar
  9. Koolbergen DR et al (2002) The pathology of fresh and cryopreserved homograft heart valves: an analysis of forty explanted homograft valves. J Thorac Cardiovasc Surg 124:689–697PubMedCrossRefGoogle Scholar
  10. Legare JF, Lee TD, Creaser K, Ross DB (2000a) T lymphocytes mediate leaflet destruction and allograft aortic valve failure in rats. Ann Thorac Surg 70:1238–1245PubMedCrossRefGoogle Scholar
  11. Legare JF, Lee DG, Ross DB (2000b) Cryopreservation of rat aortic valves results in increased structural failure. Circulation 102:III75–III78Google Scholar
  12. Lisy M, Pennecke J, Brockbank KGM et al (2010) The performance of ice-free cryopreserved heart valve allografts in an orthotopic pulmonary sheep model. Biomaterials 31:5306–5311PubMedCrossRefGoogle Scholar
  13. O’Brien MF et al (2001) The homograft aortic valve: a 29-year, 99.3% follow up of 1, 022 valve replacements. J Heart Valve Dis 10:334–344PubMedGoogle Scholar
  14. Pearson K, Dock N, Brubaker S (eds) (2008) Standards for tissue banking, 12th edn. American Association of Tissue Banks, McLean, VA, USAGoogle Scholar
  15. Rajani B, Mee RB, Ratliff NB (1998) Evidence for rejection of homograft cardiac valves in infants. J Thorac Cardiovasc Surg 115:111–117PubMedCrossRefGoogle Scholar
  16. Schenke-Layland K et al (2006) Impact of cryopreservation on extracellular matrix structures of heart valve leaflets. Ann Thorac Surg 81:918–926PubMedCrossRefGoogle Scholar
  17. Schenke-Layland K et al (2007) Optimized preservation of extracellular matrix in cryopreserved heart valves—implications for long-term durability. Ann Thorac Surg 83:1641–1650PubMedCrossRefGoogle Scholar
  18. Song YC, Khirabadi BS, Lightfoot FG, Brockbank KGM, Taylor MJ (2000) Vitreous cryopreservation maintains the function of vascular grafts. Nat Biotech 18:296–299CrossRefGoogle Scholar
  19. Stock UA et al (2000) Tissue engineered three leaflet heart valves. J Thorac Cardiovasc Surg 119:732–740PubMedCrossRefGoogle Scholar
  20. Watts LK, Duffy P, Field RB, Stafford EG, O’Brien MF (1976) Establishment of a viable homograft cardiac valve bank: a rapid method of determining homograft viability. Ann Thorac Surg 21:230–236PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Kelvin G. M. Brockbank
    • 1
    • 2
    • 3
    Email author
  • Katja Schenke-Layland
    • 4
    • 5
    • 6
  • Elizabeth D. Greene
    • 1
  • Zhenzhen Chen
    • 1
  • Olaf Fritze
    • 7
  • Martina Schleicher
    • 7
  • Renate Kaulitz
    • 8
  • Iris Riemann
    • 9
  • Falko Fend
    • 10
  • Johannes M. Albes
    • 11
  • Ulrich A. Stock
    • 7
  • Milan Lisy
    • 7
  1. 1.Cell and Tissue Systems, Inc.North CharlestonUSA
  2. 2.Institute for Bioengineering and BioscienceGeorgia Institute of TechnologyAtlantaUSA
  3. 3.Deparment of Regenerative Medicine and Cell BiologyMedical University of South CarolinaCharlestonUSA
  4. 4.Cardiovascular Research LaboratoriesDavid Geffen School of Medicine at UCLALos AngelesUSA
  5. 5.Deparment of Cell and Tissue EngineeringFraunhofer Institute for Interfacial Engineering and Biotechnology (IGB)StuttgartGermany
  6. 6.Inter-University Centre for Medical Technology Stuttgart-Tübingen (IZST)Eberhard Karls University TübingenTübingenGermany
  7. 7.Deparment of Thoracic, Cardiac and Vascular SurgeryUniversity Hospital TuebingenTuebingenGermany
  8. 8.Deparment of Pediatric CardiologyUniversity Hospital TuebingenTuebingenGermany
  9. 9.Fraunhofer Institute of Biomedical TechnologySt. IngbertGermany
  10. 10.Deparment of General Pathologie and Pathological AnatomyUniversity Hospital TuebingenTuebingenGermany
  11. 11.Deparment of Cardiac SurgeryHeart Centre BrandenburgBernauGermany

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