Cai, X

Markus, PM

Roher, HD

Selvaggi, G

Starzl, TE

English

PubMed

Name not available

Tolerance to skin and vascularized cardiac allografts using mixed chimerism

D-Scholarship@Pitt

i , , \ i II Tolerance to Skin and Vascularized Cardiac Allografts Using Mixed Chimerism P.M. Markus, G. Selvaggi, X. Cai, T.E. Starzl, and H.D. Roher I N recent years a number of methods have been devel-oped to induce donor-specific transplantation tolerance in adult recipients using bone marrow transplantation. 1-4 One approach. using a mixture of syngeneic and allogeneic bone marrow to prepare allogeneic chimeras (A + B - A) has been demonstrated to be effective in achieving long-term donor-specific transplantation tolerance to alloanti-gens in mice. 5 We adapted the model of mixed chimerism % Grafts Surviving 100 ~K------__ 80" 20., Normal LEWIS Syng.BMTX (LEW ->LEW) • MIXED BMTX (LEW+ACI ->LEW) Syngeneic Repopulation MIXED BMTX (LEW+ACI->LEW) Mixed Allogeneic Repopulation o~ •• ~[---------~----------~-a 15 30 45 60 75 90 105 120 135 150 DAYS POST ACI SKIN GRAFT % Grafts Surviving lMM~~f----------------80., 60-: ::: , i:! 401 ! 11'-20, , , [ Normal LEWIS <t Syng.BMTX (LEW ->LEW) • MIXED BMTX (LEW+ACI ->LEW) Syngeneic Repopulation MIXED BMTX (LEW+ACI->LEW) Mixed Allogeneic Repopulation o -4-~D •• h-i -----,-----------------15 30 45 60 75 90 105 120 135 150 b DAYS POST ACI CARDIAC GRAFT Fig 1. Life table survival of ACI skin (A) or cardiac (B) grafts in naive, syngeneic, reconstiMed, and mixed chimeras. Four weeks after bone marrow transplantation, each animal received an ACI, LEW, and BN skin, or an ACI cardiac graft (only donor type ACI grafts are presented). Animals that received mixed bone marrow yet repopulated syngeneically were analyzed separately (MIXED BMTX, syngeneic repopulation). Each group consisted of six to 10 animals. to the rat to investigate the response of mixed allogeneic chimeras to skin and primarily vascularized organ grafts that are limited in mice due to technical difficulties. MATERIALS AND METHODS Bone Marrow Transplantation LEW recipient rats (225 to 250 g) were lethally irradiated with 1000 rad total body irradiation and reconstituted with a mixture of syngeneic LEW (7 x 106 cells/recipients) and allogeneic ACI (43 x 106 cells/recipient) T cell-depleted bone marrow (LEW + ACI-LEW1. T-cell depletion (TCD) was performed using mag-netic polymer beads (Dynabeads M4501. Bioproducts for Science. IndianapolIs. lndl coated with monoclonal antibody (MAbl OX-19 (anti-CDS). Flow cytometnc analyses of spleen controls were routinely pertormed to assess successful depletion of OX-19+ T cells. Syngeneic reconstituted controls received 50 x 100 TCD syngeneic bone marrow cells (LEW-LEW). Assessment of Chimerism Four weeks after reconstitution with mixed allogeneic bone mar-row. animals were assessed for the presence of allogeneic donor type ACI T cells. Strain-specific MAb for the RTI.A major histocompatibility complex (MHO class I antigen was used as primary biotinylated antibody. Organ Grafting Full thickness skin grafts of donor (ACl). third party (BN). and host type (LEW) were performed using a modified method of Billingham.6 A heterotopic abdominal heart transplant model was used for vascularized grafts. RESULTS Survival of Mixed Allogeneic Reconstituted Rats Recipient survival after transplantation of the mixed bone marrow inoculum was 96% at 150 days. Thirty seven percent of all animals (19 of 52) repopulated as mixed chimeras with detectable ACI donor type peripheral blood lymphocytes. The remainder repopulated syngeneically, From the Department of Surgery, University of Dusseldorf, Dusseldorf, Germany (P.M.M., H.D.R.), and the Department of Surgery, Division of Transplantation Surgery, University of Pitts-burgh, Pittsburgh, Pennsylvania (G.S., X.C., T.E.S.). Supported by Research Grant DK 29961 from the National Institutes of Health, Bethesda. Md, the Veterans Administration, and the Deutsche Forschungsgemeinschaft. Address reprint requests to Peter M. Markus. MD. Department of Surgery, University of Dusseldorf, Moorenstrasse 5, 4000 Dusseldorf 1, Germany. .:9 1992 by Appleton & Lange 0041-1345/921$3.00/ +0 2894 Transplantation Proceedings, Vol 24, No 6 (December), 1992: pp 2894-2895 Il JL.r::nJ"\n\.,r:;: TO SKIN despite the mixed ACI + LEW bone marrow transplants. Subsequently, the animals that repopulated as mixed chi-meras were used for further investigation (I) to assess the . percentage of chimerism and (2) for transplantation exper-iments with skin and cardiac iso- and allografts. Animals that repopulated as mixed chimeras were evalu-ated at I, 3. and 5 months after reconstitution for the percentage of chimerism. The percentage of donor type allogeneic ACI cells varied from 0.5 to 82% and this was maintained throughout the study. Survival of Skin and Cardiac Grafts in Mixed Allogeneic Reconstituted Rats Although syngeneic repopulated animals rejected donor (AC!) and third-party (BN) skin grafts. all chimeric ani-mals accepted host (LEW) (data not shown) and donor type (AC!) (Fig la and b) skin and cardiac grafts. but rt:Jected third-party (BN) grafts at the same time as naive controls (data not shown). The level of chimerism did not influence the survival of skin or cardiac grafts. All donor type skin and cardiac grafts survived more than 150 days without histologic evidence of chronic rejection. Animals that repopulated syngeneically without detectable chimer-ism uniformly rejected ACI and BN skin and cardiac grafts. DISCUSSION We report a model that achieves stable mixed allogeneic chimerism using transplantation of a mixture of TCD 2895 allogeneic and TCD syngeneic bone marrow. Transplanta-tion tolerance was specific for skin and also vascularized cardiac grafts across an MHC and non-MHC barrier (ACI-LEW). The model is still limited by a high number of rats rejecting the allogeneic bone marrow inoculum and repopulating syngeneically. Neither increasing the total number of allogeneic cells nor increasing the number of totally transplanted cells up to 200 x 10° cells/recipient (data not shown) promoted the acceptance of donor type ACI marrow. It is well recognized that TCD of the syngeneic component of the mixed bone marrow inoculum is critical in order to allow engraftment of the allogeneic bone marrow cells when mixed allogeneic reconstitution is performed.7 One could argue that TCD using OX-I9-coated magnetic beads is inefficient in removing all synge-neic CD-5+ T cells. which then prevents engraftment of the ACI bone marrow component. It is also possible that the rat exhibits a greater level of alloresistance to engraft-ment that we have found in mouse-to-rat and rat-to-mouse transplantation experiments (data not shown). REFERENCES 1. Rapaport FT. et a1: Transplantation Proc 9:891. 1977 2. Slavin S. Strober R. Fuks Z. et al: 1 Exp Med 146:34. 1977 3. Dittmer J. Bennett M: Molec Cell Biochem 21:83.1978 4. Myburgh lA. Smit lA. Browde S. et a1: Transplantation 29:401, 1980 5. Ildstad ST. Sachs DH: Nature 307: 168. 1984 6. Billingham RE: In: Billingham RE. Silvers WK (eds). Trans-plantation of Tissues and Cells. Philadelphia: Wistar Institute Press. 1961. p I 7. Ildstad ST. Wren SM. Bluestone lA. et al: J Immunol 136:28. 1986 

Markus, P M

Selvaggi, G

Cai, X

Starzl, T E

Roher, H D

University of Miami: Scholarship Miami

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Tolerance to skin and vascularized cardiac allografts using mixed chimerism

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