Skip to main content

Advertisement

SpringerLink
Log in

Xenogeneic transplantation of articular chondrocytes into full-thickness articular cartilage defects in minipigs: fate of cells and the role of macrophages

  • Regular Article
  • Published:
Cell and Tissue Research Aims and scope Submit manuscript

Abstract

Xenogeneic or allogeneic chondrocytes hold great potential to build up new cartilage in vivo. However, immune rejection is a major concern for the utility of universal donor-derived cells. In order to verify the reported immune privilege of chondrocytes in vivo, the aim of this study was to assess engraftment of human articular chondrocytes (HAC) in minipig knee cartilage defects and their contribution to cartilage regeneration. HAC were transplanted matrix-assisted within two hydrogels into full-thickness cartilage defects of minipigs or implanted ectopically into immune deficient mice to assess redifferentiation capacity. At 2 and 4 weeks after surgery, cell-persistence and host cell invasion were monitored by species-specific in situ hybridization and RT-PCR. Early tissue regeneration was evaluated by histomorphometry and a modified O’Driscoll score. HAC capable of successful in vivo chondrogenic redifferentiation persisted at ectopic sites for 4 weeks in both carrier materials. Early defect regeneration involved extensive host cell invasion and a decline of HAC to less than 5 % of initial cell numbers in 6/12 defects within 2 weeks. Few clusters of persisting HAC within collagen type II-rich tissue were surrounded by porcine macrophages. Four weeks after cell transplantation, most of the defects contained well-integrated cell-rich tissue free of human cells with no apparent difference between hydrogel carriers. In summary, HAC failed to engraft in porcine articular cartilage defects despite their ability for successful in vivo redifferentiation. The co-localization of macrophages to hydrogel-implanted HAC suggests active graft rejection without evidence for an immune-privileged status of xenogeneic chondrocytes in a large animal joint.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Abe M, Cheng J, Qi J, Glaser RM, Thall AD, Sykes M, Yang YG (2002) Elimination of porcine hemopoietic cells by macrophages in mice. J Immunol 168:621–628

    Article  CAS  PubMed  Google Scholar 

  • Adkisson HD, Milliman C, Zhang X, Mauch K, Maziarz RT, Streeter PR (2010) Immune evasion by neocartilage-derived chondrocytes: Implications for biologic repair of joint articular cartilage. Stem Cell Res 4:57–68

    Article  CAS  PubMed  Google Scholar 

  • Blazar BR, Lindberg FP, Ingulli E, Panoskaltsis-Mortari A, Oldenborg PA, Iizuka K, Yokoyama WM, Taylor PA (2001) Cd47 (integrin-associated protein) engagement of dendritic cell and macrophage counterreceptors is required to prevent the clearance of donor lymphohematopoietic cells. J Exp Med 194:541–549

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Brittberg M, Lindahl A, Nilsson A, Ohlsson C, Isaksson O, Peterson L (1994) Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation. N Engl J Med 331:889–895

    Article  CAS  PubMed  Google Scholar 

  • Cadili A, Kneteman N (2008) The role of macrophages in xenograft rejection. Transplant Proc 40:3289–3293

    Article  CAS  PubMed  Google Scholar 

  • Caplan AI, Dennis JE (2006) Mesenchymal stem cells as trophic mediators. J Cell Biochem 98:1076–1084

    Article  CAS  PubMed  Google Scholar 

  • Chahal J, Gross AE, Gross C, Mall N, Dwyer T, Chahal A, Whelan DB, Cole BJ (2013) Outcomes of osteochondral allograft transplantation in the knee. Arthroscopy 29:575–588

    Article  PubMed  Google Scholar 

  • Chesterman PJ, Smith AU (1968) Homotransplantation of articular cartilage and isolated chondrocytes. An experimental study in rabbits. J Bone Joint Surg (Br) 50:184–197

    CAS  Google Scholar 

  • Dell’Accio F, De Bari C, Luyten FP (2001) Molecular markers predictive of the capacity of expanded human articular chondrocytes to form stable cartilage in vivo. Arthritis Rheum 44:1608–1619

    Article  PubMed  Google Scholar 

  • Domm C, Fay J, Schunke M, Kurz B (2000) redifferentiation of dedifferentiated joint cartilage cells in alginate culture. Effect of intermittent hydrostatic pressure and low oxygen partial pressure. Orthopade 29:91–99

    CAS  PubMed  Google Scholar 

  • Elves MW, Zervas J (1974) An investigation into the immunogenicity of various components of osteoarticular grafts. Br J Exp Pathol 55:344–351

    CAS  PubMed Central  PubMed  Google Scholar 

  • Flavell DJ, Jones DB, Wright DH (1987) Identification of tissue histiocytes on paraffin sections by a new monoclonal antibody. J Histochem Cytochem 35:1217–1226

    Article  CAS  PubMed  Google Scholar 

  • Glant T, Mikecz K (1986) Antigenic profiles of human, bovine and canine articular chondrocytes. Cell Tissue Res 244:359–369

    CAS  PubMed  Google Scholar 

  • Goebel U, Mecklenburg A, Siepe M, Roesslein M, Schwer CI, Pahl HL, Priebe HJ, Schlensak C, Loop T (2009) Protective effects of inhaled carbon monoxide in pig lungs during cardiopulmonary bypass are mediated via an induction of the heat shock response. Br J Anaesth 103:173–184

    Article  CAS  PubMed  Google Scholar 

  • Gotterbarm T, Richter W, Jung M, Berardi Vilei S, Mainil-Varlet P, Yamashita T, Breusch SJ (2006) An in vivo study of a growth-factor enhanced, cell free, two-layered collagen-tricalcium phosphate in deep osteochondral defects. Biomaterials 27:3387–3395

    Article  CAS  PubMed  Google Scholar 

  • Hirschmann F, Verhoeyen E, Wirth D, Bauwens S, Hauser H, Rudert M (2002) Vital marking of articular chondrocytes by retroviral infection using green fluorescence protein. Osteoarthr Cartil 10:109–118

    Article  CAS  PubMed  Google Scholar 

  • Hoemann CD (2011) International cartilage repair society (icrs) recommended guidlines for histological endpoints for cartilage repair studies in animal models and clinical trials. Cartilage 2:153–172

    Article  CAS  Google Scholar 

  • Huey DJ, Sanchez-Adams J, Willard VP, Athanasiou KA (2012) Immunogenicity of bovine and leporine articular chondrocytes and meniscus cells. Tissue Eng A 18:568–575

    Article  CAS  Google Scholar 

  • Hunt HE, Sadr K, Deyoung AJ, Gortz S, Bugbee WD (2014) The role of immunologic response in fresh osteochondral allografting of the knee. Am J Sports Med 42:886–891

    Article  PubMed  Google Scholar 

  • Ide K, Ohdan H, Kobayashi T, Hara H, Ishiyama K, Asahara T (2005) Antibody- and complement-independent phagocytotic and cytolytic activities of human macrophages toward porcine cells. Xenotransplantation 12:181–188

    Article  PubMed  Google Scholar 

  • Ide K, Wang H, Tahara H, Liu J, Wang X, Asahara T, Sykes M, Yang YG, Ohdan H (2007) Role for cd47-sirpalpha signaling in xenograft rejection by macrophages. Proc Natl Acad Sci U S A 104:5062–5066

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Jobanputra P, Corrigall V, Kingsley G, Panayi G (1992) Cellular responses to human chondrocytes: Absence of allogeneic responses in the presence of hla-dr and icam-1. Clin Exp Immunol 90:336–344

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Jung M, Kaszap B, Redohl A, Steck E, Breusch S, Richter W, Gotterbarm T (2009) Enhanced early tissue regeneration after matrix-assisted autologous mesenchymal stem cell transplantation in full thickness chondral defects in a minipig model. Cell Transplant 18:923–932

    Article  PubMed  Google Scholar 

  • Kasten P, Vogel J, Luginbuhl R, Niemeyer P, Tonak M, Lorenz H, Helbig L, Weiss S, Fellenberg J, Leo A, Simank HG, Richter W (2005) Ectopic bone formation associated with mesenchymal stem cells in a resorbable calcium deficient hydroxyapatite carrier. Biomaterials 26:5879–5889

    Article  CAS  PubMed  Google Scholar 

  • Krase A, Steck E, Roth W, Richter W (2013) Biomechanics of cartilage tissue engineering constructs : Sensitive test procedure for assessment of biomechanical functionality and further development after in vivo transplantation. Orthopade 42:262–270

    Article  CAS  PubMed  Google Scholar 

  • Krase A, Abedian R, Steck E, Hurschler C, Richter W (2014) Bmp activation and wnt-signalling affect biochemistry and functional biomechanical properties of cartilage tissue engineering constructs. Osteoarthr Cartil 22:284–292

    Article  CAS  PubMed  Google Scholar 

  • Kuhne M, Erben U, Schulze-Tanzil G, Kohler D, Wu P, Richter FJ, John T, Radbruch A, Sieper J, Appel H (2009) Hla-b27-restricted antigen presentation by human chondrocytes to cd8+ t cells: Potential contribution to local immunopathologic processes in ankylosing spondylitis. Arthritis Rheum 60:1635–1646

    Article  CAS  PubMed  Google Scholar 

  • Kurz B, Lemke AK, Fay J, Pufe T, Grodzinsky AJ, Schunke M (2005) Pathomechanisms of cartilage destruction by mechanical injury. Ann Anat 187:473–485

    Article  CAS  PubMed  Google Scholar 

  • Langer F, Gross AE (1974) Immunogenicity of allograft articular cartilage. J Bone Joint Surg Am 56:297–304

    CAS  PubMed  Google Scholar 

  • Levy YD, Gortz S, Pulido PA, McCauley JC, Bugbee WD (2013) Do fresh osteochondral allografts successfully treat femoral condyle lesions? Clin Orthop Relat Res 471:231–237

    Article  PubMed Central  PubMed  Google Scholar 

  • Mantovani A, Biswas SK, Galdiero MR, Sica A, Locati M (2013) Macrophage plasticity and polarization in tissue repair and remodelling. J Pathol 229:176–185

    Article  CAS  PubMed  Google Scholar 

  • Masuoka K, Asazuma T, Ishihara M, Sato M, Hattori H, Ishihara M, Yoshihara Y, Matsui T, Takase B, Kikuchi M, Nemoto K (2005) Tissue engineering of articular cartilage using an allograft of cultured chondrocytes in a membrane-sealed atelocollagen honeycomb-shaped scaffold (achms scaffold). J Biomed Mater Res B 75:177–184

    Article  Google Scholar 

  • Matricali GA, Dereymaeker GP, Luyten FP (2010) Donor site morbidity after articular cartilage repair procedures: A review. Acta Orthop Belg 76:669–674

    PubMed  Google Scholar 

  • McCulloch PC, Kang RW, Sobhy MH, Hayden JK, Cole BJ (2007) Prospective evaluation of prolonged fresh osteochondral allograft transplantation of the femoral condyle: Minimum 2-year follow-up. Am J Sports Med 35:411–420

    Article  PubMed  Google Scholar 

  • Mierisch CM, Wilson HA, Turner MA, Milbrandt TA, Berthoux L, Hammarskjold ML, Rekosh D, Balian G, Diduch DR (2003) Chondrocyte transplantation into articular cartilage defects with use of calcium alginate: The fate of the cells. J Bone Joint Surg Am 85-A:1757–1767

    PubMed  Google Scholar 

  • Navarro-Alvarez N, Yang YG (2014) Lack of cd47 on donor hepatocytes promotes innate immune cell activation and graft loss: A potential barrier to hepatocyte xenotransplantation. Cell Transplant 23:345–354

    Article  PubMed Central  PubMed  Google Scholar 

  • Oldenborg PA, Zheleznyak A, Fang YF, Lagenaur CF, Gresham HD, Lindberg FP (2000) Role of cd47 as a marker of self on red blood cells. Science 288:2051–2054

    Article  CAS  PubMed  Google Scholar 

  • Ostrander RV, Goomer RS, Tontz WL, Khatod M, Harwood FL, Maris TM, Amiel D (2001) Donor cell fate in tissue engineering for articular cartilage repair. Clin Orthop Relat Res. 389:228–237

  • Pelttari K, Lorenz H, Boeuf S, Templin MF, Bischel O, Goetzke K, Hsu HY, Steck E, Richter W (2008) Secretion of matrix metalloproteinase 3 by expanded articular chondrocytes as a predictor of ectopic cartilage formation capacity in vivo. Arthritis Rheum 58:467–474

    Article  CAS  PubMed  Google Scholar 

  • Rahfoth B, Weisser J, Sternkopf F, Aigner T, von der Mark K, Brauer R (1998) Transplantation of allograft chondrocytes embedded in agarose gel into cartilage defects of rabbits. Osteoarthr Cartil 6:50–65

    Article  CAS  PubMed  Google Scholar 

  • Ramallal M, Maneiro E, Lopez E, Fuentes-Boquete I, Lopez-Armada MJ, Fernandez-Sueiro JL, Galdo F, De Toro FJ, Blanco FJ (2004) Xeno-implantation of pig chondrocytes into rabbit to treat localized articular cartilage defects: An animal model. Wound Repair Regen 12:337–345

    Article  PubMed  Google Scholar 

  • Revell CM, Athanasiou KA (2009) Success rates and immunologic responses of autogenic, allogenic, and xenogenic treatments to repair articular cartilage defects. Tissue Eng B 15:1–15

    Article  CAS  Google Scholar 

  • Romaniuk A, Malejczyk J, Kubicka U, Hyc A, Olszewski WL, Moskalewski S (1995) Rejection of cartilage formed by transplanted allogeneic chondrocytes: Evaluation with monoclonal antibodies. Transpl Immunol 3:251–257

    Article  CAS  PubMed  Google Scholar 

  • Schulze-Tanzil G, de Souza P, Villegas Castrejon H, John T, Merker HJ, Scheid A, Shakibaei M (2002) Redifferentiation of dedifferentiated human chondrocytes in high-density cultures. Cell Tissue Res 308:371–379

    Article  CAS  PubMed  Google Scholar 

  • Steck E, Burkhardt M, Ehrlich H, Richter W (2010) Discrimination between cells of murine and human origin in xenotransplants by species specific genomic in situ hybridization. Xenotransplantation 17:153–159

    Article  PubMed  Google Scholar 

  • van Susante JL, Buma P, Schuman L, Homminga GN, van den Berg WB, Veth RP (1999) Resurfacing potential of heterologous chondrocytes suspended in fibrin glue in large full-thickness defects of femoral articular cartilage: An experimental study in the goat. Biomaterials 20:1167–1175

    Article  PubMed  Google Scholar 

  • Wu G, Korsgren O, Zhang J, Song Z, van Rooijen N, Tibell A (2000) Pig islet xenograft rejection is markedly delayed in macrophage-depleted mice: A study in streptozotocin diabetic animals. Xenotransplantation 7:214–220

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The study was funded by the Federal Ministry of Education and Research (BMBF grants 0315577H and 0315579) as well as a grant from TETEC AG, Reutlingen, Germany. We thank Jürgen Mollenhauer for providing porcine albumin for Novocart® Inject preparation. We gratefully acknowledge Elisabeth Seebach, Martin Rütze and Jan Brocher for help and advice and Nicole Buchta and Birgit Frey for excellent technical assistance.

Disclosure of conflicts

W. Richter received financial support for conducting this study from TETEC AG, Reutlingen, Germany. TETEC AG had no influence on data analysis and the manuscript writing of this work.

Author information

Authors and Affiliations

  1. Research Center for Experimental Orthopaedics, Orthopaedic University Hospital Heidelberg, Schlierbacher Landstrasse 200a, 69118, Heidelberg, Germany

    Thomas Niemietz, Gesa Zass, Solvig Diederichs & Wiltrud Richter

  2. Clinic for Orthopaedics and Trauma Surgery, Orthopaedic University Hospital Heidelberg, Schlierbacher Landstr. 200a, 69118, Heidelberg, Germany

    Sébastien Hagmann & Tobias Gotterbarm

Authors
  1. Thomas Niemietz
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Gesa Zass
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sébastien Hagmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Solvig Diederichs
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Tobias Gotterbarm
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Wiltrud Richter
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Wiltrud Richter.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Fig. 1

Species-specific genomic in situ hybridization. Expanded HAC embedded in fibrin were cultured under chondrogenic conditions for 14 days to obtain a stable construct with cartilaginous matrix deposition. Then, the sample was coated by a layer of expanded porcine mesenchymal stromal cells embedded in fibrin and cultured for further 5 days. Serial sections of the composites were subjected to species-specific in situ hybridization for simultaneous detection of a human cells (ALU DNA repeats) and b porcine cells (pSINE hybridization). A dark signal indicates positive cell nuclei. The letters mark areas with human (h) or porcine (p) cell layers. No cross-hybridization to cells of the other species was evident (GIF 144 kb)

Supplementary Fig. 1

High resolution image file (TIF 13.7 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Niemietz, T., Zass, G., Hagmann, S. et al. Xenogeneic transplantation of articular chondrocytes into full-thickness articular cartilage defects in minipigs: fate of cells and the role of macrophages. Cell Tissue Res 358, 749–761 (2014). https://doi.org/10.1007/s00441-014-1982-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00441-014-1982-x

Keywords

Search

Navigation