Nuclear transplantation (therapeutic cloning) could theoretically provide a limitless source of cells for regenerative therapy. Although the cloned cells would carry the nuclear genome of the patient, the presence of mitochondria inherited from the recipient oocyte raises questions about the histocompatibility of the resulting cells. In this study, we created bioengineered tissues from cardiac, skeletal muscle, and renal cells cloned from adult bovine fibroblasts. Long-term viability was demonstrated after transplantation of the grafts into the nuclear donor animals. Reverse transcription-PCR (RT-PCR) and western blot analysis confirmed that the cloned tissues expressed tissue-specific mRNA and proteins while expressing a different mitochondrial DNA (mtDNA) haplotype. In addition to creating skeletal muscle and cardiac “patches”, nuclear transplantation was used to generate functioning renal units that produced urinelike fluid and demonstrated unidirectional secretion and concentration of urea nitrogen and creatinine. Examination of the explanted renal devices revealed formation of organized glomeruli- and tubule-like structures. Delayed-type hypersensitivity (DTH) testing in vivo and Elispot analysis in vitro suggested that there was no rejection response to the cloned renal cells. The ability to generate histocompatible cells using cloning techniques addresses one of the major challenges in transplantation medicine.
This is a preview of subscription content
Subscribe to Journal
Get full journal access for 1 year
only $8.25 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
Lanza, R.P. et al. The ethical reasons for stem cell research. Science 293, 1299 (2001).
Atala, A. & Lanza, R.P. Methods of Tissue Engineering (Academic Press, San Diego, CA, 2001).
Atala, A. & Mooney, D. Synthetic Biodegradable Polymer Scaffolds (Birkhaüser, Boston, MA, 1997).
Machluf, M. & Atala, A. Emerging concepts for tissue and organ transplantation. Graft 1, 31–37 (1998).
Lanza, R.P., Cibelli, J.B. & West, M.D. Prospects for the use of nuclear transfer in human transplantation. Nat. Biotechnol. 17, 1171–1174 (1999).
Evans, M.J. et al. Mitochondrial DNA genotypes in nuclear transfer-derived cloned sheep. Nat. Genet. 23, 90–93 (1999).
Hiendleder, S., Schmutz, S.M., Erhardt, G., Green, R.D. & Plante, Y. Transmitochondrial differences and varying levels of heteroplasmy in nuclear transfer cloned cattle. Mol. Reprod. Dev. 54, 24–31 (1999).
Steinborn, R. et al. Mitochondrial DNA heteroplasmy in cloned cattle produced by fetal and adult cell cloning. Nat. Genet. 25, 255–257 (2000).
Vyas, J.M. et al. Biochemical specificity of H-2M3a: stereospecificity and space-filling requirement at position 1 maintains N-formyl peptide binding. J. Immunol. 149, 3605–3611 (1992).
Morse, M. et al. The COI mitochondrial gene encodes a minor histocompatibility antigen presented by H2-M3. J. Immunol. 156, 3301–3307 (1996).
Loveland, B., Wang, C.R., Yonekawa, H., Hermel, E. & Lindahl, K.F. Maternally transmitted histocompatibility antigens of mice: a hydrophobic peptide of a mitochondrial encoded protein. Cell 60, 971–980 (1990).
Davies, J.D. et al. Generation of T cells with lytic specificity for atypical antigens. I. A mitochondrial antigen in the rat. J. Exp. Med. 173, 823–832 (1991).
Lysaght, M.J. Maintenance dialysis population dynamics: current trends and long-term implications. J. Am. Soc. Nephrol. 13, S37–S40 (2002).
Amiel, G.E. & Atala, A. Current and future modalities for functional renal replacement. Urol. Clin. 26, 235–246 (1999).
Humes, H.D., Buffington, D.A., MacKay, S.M., Funke, A.J. & Weitzel, W.F. Replacement of renal function in uremic animals with a tissue-engineered kidney. Nat. Biotechnol. 17, 451–455 (1999).
Cieslinski, D.A. & Humes, H.D. Tissue engineering of a bioartificial kidney. Biotechnol. Bioeng. 43, 781–791 (1994).
MacKay, S.M., Kunke, A.J., Buffington, D.A. & Humes, H.D. Tissue engineering of a bioartificial renal tubule. ASAIO J. 44, 179–183 (1998).
Aebischer, P., Ip, T.K., Panol, G. & Galletti, P.M. The bioartificial kidney: progress towards an ultrafiltration device with renal epithelial cells processing. Life Support Syst. 5, 159–168 (1987).
Ip, T., Aebischer, P. & Galletti, P.M. Cellular control of membrane permeability. Implications for a bioartificial renal tubule. ASAIO Trans. 34, 351–355 (1988).
Humes, H.D. Renal replacement devices. in Principles of Tissue Engineering; Edn. 2 (eds Lanza, R.P., Langer, R. & Vacanti, J.) 645–653 (Academic Press, San Diego, 2000).
Amiel, A., Yoo, J. & Atala, A. Renal therapy using tissue engineered constructs and gene delivery. World J. Urol. 18, 71–79 (2000).
Lanza, R.P., Hayes, J.L. & Chick, W.L. Encapsulated cell technology. Nat. Biotechnol. 14, 1107–1111 (1996).
Kuhtreiber, W.M., Lanza, R.P. & Chick, W.L. (eds). Cell Encapsulation Technology and Therapeutics (Birkhauser, Boston, 1998).
Lanza, R.P & Chick, W.L. (eds). Immunoisolation of Pancreatic Islets (R.G. Landes, Austin, TX, 1994).
Joki, T. et al. Continuous release of endostatin from microencapsulated engineered cells for tumor therapy. Nat. Biotechnol. 19, 35–39 (2001).
Qiao, J., Sakurai, H. & Nigam, S.K. Branching morphogenesis independent of mesenchymal-epithelial contact in the developing kidney. Proc. Natl. Acad. Sci. USA 96, 7330–7335 (1999).
Humes, H.D., Krauss, J.C., Cieslinski, D.A. & Funke, A.J. Tubulogenesis from isolated single cells of adult mammalian kidney: clonal analysis with a recombinant retrovirus. Am. J. Physiol. 271, F42–F49 (1996).
Lanza, R.P, Langer, R. & Vacanti, J. Principles of Tissue Engineering (Academic Press, San Diego, CA, 2000).
Atala, A. Future perspectives in reconstructive surgery using tissue engineering. Urol. Clin. 26, 157–166 (1999).
Santavirta, S. et al. Immune response to polyglycolic acid implants. J. Bone Joint Surg. Br. 72, 597–600 (1990).
Paivarinta, U. et al. Intraosseous cellular response to biodegradable fracture fixation screws made of polyglycolide or polylactide. Arch. Orthop. Trauma Surg. 112, 71–74 (1993).
Bostman, O.M. & Pihlajamaki, H.K. Adverse tissue reactions to bioabsorbable fixation devices. Clin. Orthop. 371, 216–227 (2000).
Ruuskanen, M. et al. Evaluation of self-reinforced polyglycolide membrane implanted in the subcutis of rabbits. Ann. Chir. Gynaecol. 88 308–312 (1999).
Weiler, A., Helling, H.J., Kirch, U., Zirbes, T.K. & Rehm, K.E. Foreign-body fracture fixation: experimental study in sheep. J. Bone Joint Surg. Br. 78, 369–376 (1996).
Pariente, J.L., Kim, B.S. & Atala, A. In vitro compatibility assessment of naturally-derived and synthetic biomaterials using normal human urothelial cells. J. Biomed. Mat. Res. 55, 33–39 (2001).
Rosenberger, G. Clinical Examination of Cattle (Verlag Paul Parey, Berlin, 1979), pp.275–281.
Smith, B.P. Large Animal Internal Medicine: Diseases of Horses, Cattle, Sheep and Goats, Edn. 2 pp. 467–469 (Mosby, St. Louis, 1996).
Lanza, R.P. et al. Cloning of an endangered species (Bos gaurus) using interspecies nuclear transfer. Cloning 2, 79–90 (2000).
Fischer Lindahl, K., Hermel, E., Loveland, B.E. & Wang, C.R. Maternally transmitted antigen of mice. Ann. Rev. Immunol. 9, 351–372 (1991).
Hadley, G.A., Linders, B. & Mohanakumar, T. Immunogenicity of MHC class I alloantigens expressed on parenchymal cells in the human kidney. Transplantation 54, 537–542 (1992).
Yard, B.A. et al. Analysis of T cell lines from rejecting renal allografts. Kidney Int. 43, S133–S138 (1993).
Bailey, D.W. Genetics of histocompatibility in mice. I. New loci and congenic lines. Immunogenetics 2, 249–256 (1975).
Mohanakumar, T. The Role of MHC and Non-MHC Antigens in Allograft Immunity pp. 1–115 (R.G. Landes Company, Austin, TX, 1994).
Lanza, R.P., Cibelli, J.B. & West, M.D. Human therapeutic cloning. Nat. Med. 5, 975–977 (1999).
Cibelli, J.B. et al. Somatic cell nuclear transfer in humans: pronuclear and early embryonic development. e-biomed: J. Regen. Med. 2, 25–31 (2001).
Lanza, R.P. et al. The ethical validity of using nuclear transfer in human transplantation. JAMA 284, 3175–3179 (2000).
Itskovitz-Eldor, J. et al. Differentiation of human embryonic stem cells into embryoid bodies comprising the three embryonic germ layers. Mol. Med. 5, 88–95 (2000).
Schuldiner, M., Yanuka, O., Itskovitz-Eldor, J., Melton, D.A. & Benvenisty, N. Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells. Proc. Natl. Acad. Sci USA 97, 11307–11312 (2000).
Kaufman, D.S. et al. Directed differentiation of human embryonic stem cells into hematopoietic colony forming cells. Blood 94 (Suppl. 1, part 1 of 2), 34a (1999).
Reubinoff, B.E. et al. Neural progenitors from human embryonic stem cells. Nat. Biotechnol. 19, 1134–1140 (2001).
Reubinoff, B.E., Pera, M.F., Fong, C.Y., Trounson, A. & Bongso, A. Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro. Nat. Biotechnol. 18, 399–404 (2000).
Cibelli, J.B. et al. Parthenogenetic stem cells in nonhuman primates. Science 295, 819 (2002).
Oberpenning, F.O., Meng, J., Yoo, J. & Atala, A. De novo reconstitution of a functional urinary bladder by tissue engineering. Nat. Biotechnol. 17, 149–155 (1999).
Kaushal, S. et al. Circulating endothelial cells for tissue engineering of small diameter vessels. Nat. Med. 7, 1035–1040 (2001).
Presicce, G.A. & Yang, X. Parthenogenetic development of bovine oocytes matured in vitro for 24 hr and activated by ethanol and cycloheximide. Mol. Reprod. Dev. 38, 380–385 (1994).
We thank Jose B. Cibelli, Frederick F. Hess (DVM), R. T. Duby, and the Department of Veterinary Sciences, University of Massachusetts, Amherst. We also thank Kyung-Ha Kang (Brigham & Women's Hospital), Wendy Nevala (Mayo Clinic), and Maria P. Bayona-Bafaluy (University of Miami) for their help with the histologic, Elispot, and molecular analyses. This research was supported in part by National Institutes of Health grants AI-16052 (to P.J.W.) and RO1DK57260 (to A.A.).
R.P.L., C.B., and M.D.W. are employed by Advanced Cell Technology, which is pursuing therapeutic cloning.
About this article
Cite this article
Lanza, R., Chung, H., Yoo, J. et al. Generation of histocompatible tissues using nuclear transplantation. Nat Biotechnol 20, 689–696 (2002). https://doi.org/10.1038/nbt703
Stem Cell Research & Therapy (2021)
Current Transplantation Reports (2021)
Simple 3D culture of dissociated kidney mesenchyme mimics nephron progenitor niche and facilitates nephrogenesis Wnt-independently
Scientific Reports (2019)