Centre for Transplant and Renal Research
Westmead millennium institute
Tracking human regulatory T cells (Treg) in a humanized mouse model of porcine islet xenotransplantation
Fei Guo1, Feng Gao1, Wayne J Hawthorne1, Dandan Huang1, Sussan Davies1, Heather Burns1, Yixiong Tan1, Shounan Yi1, Philip O’Connell1.
1Centre for Transplant and Renal Research, Westmead Millennium Institute, Sydney, Australia
Introduction. Our previous study has shown that adoptive transfer with humanTreg protects against porcine islet xenograft rejection. However, where adoptively transferred Treg go and do they localize within the transplanted site to initiate their suppression of the xenoresponse remain to be answered.In this study we tracked human Treg in vivo homing by fluorescent imaging in porcine islet xenotransplantion and investigated the correlation of Treg xenograftrecruitment and their suppression of xenograft rejection. Methods. Ex vivo expanded human Treg were labelled with VivoTrack 680 (VT680, a near infrared fluorescent cell labelling agent with higher tissue penetration). The viability, phenotype and in vitro function of VT680-labelled Treg (VT680-Treg ) were assessed. Treg in vivo tracking was performed by transplantation of NOD-SCID IL2rγ-/- mice with neonatal porcine islet (NICC) under the capsule of right kidneys followed by adoptive transfer with human peripheral blood mononuclear cells (PBMC) alone or plus autologous VT680-Treg. Treg homing was imaged using Pearl Imager. NICC xenograft survival was analysed atpredetermined time points. Results. VT680-Treg remained unchanged viability, suppressive phenotype and in vitro function as well as high levels of inflammatory homing chemokine receptors CCR4, CCR5, CXCR3 and CXCR6 compared to unlabelled counterparts. As reported previously, human PBMC alone transferred recipients rejected NICC xenografts completely with no detectableinsulin-positive staining cells but infiltrated human CD45+ cells in the rejecting grafts within 30 days after PBMC transfer. By contrast, NICC xenografts remained intact with positive insulin staining cells in recipient mice beyond 60 days after receiving both human PBMC and VT680-Treg, and the prolonged NICC xenograft survival correlated with fluorescent imaging of the transplant site (right kidney). Moreover upregulated gene expression of inflammatory homing chemokines CCL17, CCL5, CXCL10 and CXCL16 for corresponding receptors CCR4, CCR5, CXCR3 and CXCR6, respectively was detected within the xenografts but not in the capsule tissue of left kidneys (no transplant site) from human PBMC and VT680-Treg transferred NICC recipients, suggesting the possible involvement of inflammatory homing in human Treg recruitment to inflamed site (xenograft), where they protect against NICC xenograft rejection.Conclusions.VT680-labelling of Treg suggests a potential approach to track Treg in real time to facilitate the understanding of the mechanisms underlying human Treg-mediated suppression of the xenogeneic response in islet xenoptransplantation. The importance of Treg xenograft homing in their in vivo modulation of the xenogeneic response is being under investigation.
15:30 - 17:00
|Islet Cell Transplantation - Miscellaneous||Tracking human regulatory T cells (Treg) in a humanized mouse model of porcine islet xenotransplantation||Room 111-112|