000109552 001__ 109552
000109552 005__ 20220203103013.0
000109552 037__ $$aTESIS-2022-018
000109552 041__ $$aeng
000109552 1001_ $$aAlonso Guallart, Paula
000109552 24500 $$aMixed hematopoietic chimerism and immune tolerance through bone marrow transplantation and infusion of regulatory T cells in a preclinical large animal model
000109552 260__ $$aZaragoza$$bUniversidad de Zaragoza, Prensas de la Universidad$$c2020
000109552 300__ $$a212
000109552 4900_ $$aTesis de la Universidad de Zaragoza$$v2021-333$$x2254-7606
000109552 500__ $$aPresentado: 07 01 2020
000109552 502__ $$aTesis-Univ. Zaragoza, , 2020$$bZaragoza, Universidad de Zaragoza$$c2020
000109552 506__ $$aall-rights-reserved
000109552 520__ $$aInduction of transplantation tolerance to kidney allografts has been achieved through transient mixed hematopoietic chimerism in a non-myeloablative approach in both, non-human primates and humans. In order to make this approach applicable to other organs less tolerogenic such as lung or heart, we studied an approach to induce long-term mixed chimerism (instead of transient) through bone marrow transplantation (BMT) and infusion of recipient in vitro-expanded regulatory T cells (Tregs) in a non-human primate model (Cynomolgus macaque). Immunosuppression monotherapy was discontinued shortly after BMT. Donor-recipient pairs were major-histocompatibility-complex (MHC) mismatched in order to increase the applicability of this approach. <br />First, we studied the biology of Mauritian Cynomolgus macaque (MCM) Tregs and developed five in vitro Treg expansion protocols for translational studies that included the use of artificial antigen presenting cells (aAPCs), donor peripheral blood mononuclear cells (PBMCs) or a pool from different donors of CD40L-stimulated B cells (CD40L-sBc). Tregs from all protocols suppressed the proliferation of anti-CD2CD3CD28 bead-stimulated autologous PBMCs albeit with different potencies, varying from 1:2-1:4 Treg:PBMC ratios, up to >1:32. Treg expansion varied between protocols but at least 1,000 fold expansion was achieved with all of the them, up to >7,000 folds. Reculture of cryopreserved Tregs permitted reexpansion with improved suppressive activity. Occasionally, CD8 contamination was observed and resolved by resorting. Specificity studies showed suppression of PBMCs from autologous cells, cells from the same donor used for stimulation during the Treg cultures and from a third‐party PBMC responders, suggestive of the polyclonallity of these Tregs. Similar to humans, the Treg–specific demethylated region (TSDR) within the FoxP3 locus correlated with suppressive activity and expression of FoxP3. Contrary to humans, FoxP3 expression did not correlate with CD45RA or CD127 expression.<br />We then investigated the efficacy of ex vivo expanded Tregs to promote the induction of durable mixed chimerism along with BMT. A total of ten recipients received Tregs with different doses of bone marrow (BM) and outcomes were compared to five controls that did not receive Tregs. Prolonged chimerism was observed in Treg-treated recipients that received a high BM dose with infusion of Tregs compared to those that received low-dose BMT or did not received Tregs. Graft-versus-host disease (GVHD) was observed in four recipients, two controls and two animals that received Tregs expanded with CD40L-sBc. In those animals in which prolonged chimerism was observed, a higher number of peripheral Tregs was detected in blood compared to baseline levels and in vitro, the anti-donor response was decreased, suggestive of donor tolerance. BM rejection and chimerism loss was associated with an inversion of the CD4 and CD8 ratios and an increase in the CD8 absolute counts. Cytomegalovirus (CMV) was detected in all recipients post-BMT independently of the administration of Tregs. CMV reactivation was associated with an increase in the CD8 counts and with the loss of the BM graft. Therefore, promptly antiviral treatment was stablished for an early CMV control. <br />In conclusion, Tregs were able to expand the duration of chimerism (albeit transient) when administered with high-dose BMT across MHC barriers without immunosuppression. <br />
000109552 520__ $$a<br />
000109552 521__ $$97086$$aPrograma de Doctorado en Medicina y Sanidad Animal
000109552 6531_ $$ainmunologia
000109552 6531_ $$atrasplante de organos
000109552 6531_ $$acultivo celular
000109552 700__ $$aBOLEA BAILO, ROSA$$edir.
000109552 700__ $$aDURAN-STRUUCK, RAIMON$$edir.
000109552 7102_ $$aUniversidad de Zaragoza$$b
000109552 830__ $$9498
000109552 8560_ $$fcdeurop@unizar.es
000109552 8564_ $$s4970203$$uhttps://zaguan.unizar.es/record/109552/files/TESIS-2022-018.pdf$$zTexto completo (eng)
000109552 909CO $$ooai:zaguan.unizar.es:109552$$pdriver
000109552 909co $$ptesis
000109552 9102_ $$a$$b
000109552 980__ $$aTESIS