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Immunoregulatory and Lipid Presentation Pathways Are Upregulated in Human Face Transplant Rejection

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Immunoregulatory and Lipid Presentation Pathways Are Upregulated in Human Face Transplant Rejection Immunoregulatory and lipid presentation pathways are upregulated in human face transplant rejection Thet Su Win, … , Leonardo V. Riella, Rachael Clark J Clin Invest. 2021. https://doi.org/10.1172/JCI135166. Clinical Medicine In-Press Preview Immunology Transplantation BACKGROUND. Rejection is the primary barrier to broader implementation of vascularized composite allografts (VCA), including face and limb transplants. The immunologic pathways activated in face transplant rejection have not been fully characterized. METHODS. Utilizing skin biopsies prospectively collected over nine years from seven face transplant patients, we studied rejection by gene expression profiling, histology, immunostaining and T cell receptor sequencing. RESULTS. Grade 1 rejection did not differ significantly from non-rejection, suggesting that it does not represent a pathologic state and that watchful waiting is warranted. In Grade 2, there was a balanced upregulation of both pro- inflammatory T cell activation pathways and anti-inflammatory checkpoint and immunomodulatory pathways, with a net result of no tissue injury. In Grade 3, IFNγ-driven inflammation, antigen presenting cell activation and infiltration of the skin by proliferative T cells bearing markers of antigen specific activation and cytotoxic effector molecules tipped the balance towards tissue injury. Rejection of VCA and solid organ transplants had both distinct and common features. VCA rejection was uniquely associated with upregulation of immunoregulatory genes, including SOCS1, induction of lipid antigen-presenting CD1 proteins, and infiltration by T cells predicted to recognize CD1b and CD1c. […] Find the latest version: https://jci.me/135166/pdf Immunoregulatory and lipid presentation pathways are upregulated in human face transplant rejection Thet Su Win1,2,3, William J. Crisler1,2, Beatrice Dyring-Andersen2, Rachel Lopdrup3, Jessica E. Teague2, Qian Zhan2, Victor Barrera4, Shannan Ho Sui4, Sotirios Tasigiorgos3, Naoka Murakami5, Anil Chandraker5, Stefan G. Tullius6, Bohdan Pomahac3, Leonardo V. Riella5*, Rachael A. Clark2* 1These authors contributed equally. 2Department of Dermatology, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA. 3Division of Plastic Surgery, Department of Surgery, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA. 4Bioinformatics Core, Department of Biostatistics, Harvard T.H. Chan School of Public Health, Boston, MA. 5Schuster Transplantation Research Center, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA. 6Division of Transplant Surgery, Department of Surgery, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA. Address for Correspondence *Rachael A. Clark M.D., Ph.D. Email: [email protected] Brigham and Women’s Hospital 1 EBRC 501A, 221 Longwood Avenue, Boston, MA 02115 Tel: (617) 525-8509 *Leonardo V. Riella M.D., Ph.D. Email: [email protected] Brigham and Women’s Hospital 221 Longwood Avenue, Boston, MA 02115 Tel: (617) 732-5500 Conflict of Interest The authors have declared that no conflict of interest exists. 2 Abstract BACKGROUND. Rejection is the primary barrier to broader implementation of vascularized composite allografts (VCA), including face and limb transplants. The immunologic pathways activated in face transplant rejection have not been fully characterized. METHODS. Utilizing skin biopsies prospectively collected over nine years from seven face transplant patients, we studied rejection by gene expression profiling, histology, immunostaining and T cell receptor sequencing. RESULTS. Grade 1 rejection did not differ significantly from non-rejection, suggesting that it does not represent a pathologic state. In Grade 2, there was a balanced upregulation of both pro- inflammatory T cell activation pathways and anti-inflammatory checkpoint and immunomodulatory pathways, with a net result of no tissue injury. In Grade 3, IFNγ-driven inflammation, antigen presenting cell activation and infiltration of the skin by proliferative T cells bearing markers of antigen specific activation and cytotoxicity tipped the balance towards tissue injury. Rejection of VCA and solid organ transplants had both distinct and common features. VCA rejection was uniquely associated with upregulation of immunoregulatory genes, including SOCS1, induction of lipid antigen-presenting CD1 proteins, and infiltration by T cells predicted to recognize CD1b and CD1c. CONCLUSIONS. Our findings suggest that the distinct features of VCA rejection reflect the unique immunobiology of skin and that enhancing cutaneous immunoregulatory networks may be a useful strategy in combatting rejection. 3 FUNDING. Department of Defense (W81XWH-17-1-0278; W81XWH-16-1-0647; W81XWH- 16-1-0689; W81XWH-18-1-0784; W81XWH-1-810798), American Society of Transplantation’s Transplantation and Immunology Research Network Fellowship Research Grant, Plastic Surgery Foundation Fellowship from the American Society of Plastic Surgeons, Novo Nordisk Foundation (NNF15OC0014092), the Lundbeck Foundation, the Aage Bangs Foundation, the A.P.Moller Foundation for the Advances of Medical Sciences, NIH #UL1 RR 025758. 4 Introduction At least forty human face transplants have been reported worldwide since the first procedure was performed in 2005. In the intervening years, face transplantation has proven to be an effective and life-transforming reconstructive technique (1, 2). It is estimated that approximately seven million people each year in the United States could benefit from vascularized composite allotransplants (VCA), such as face and limb transplants, to repair defects from trauma, tumor resections or limb loss (3). Despite promising functional and aesthetic restoration, VCA has yet to reach wide implementation in large part because of the challenges associated with diagnosing and treating rejection. VCA procedures transfer large areas of donor skin that are highly populated by donor-derived professional antigen presenting cells (APCs), including Langerhans cells, dermal dendritic cells and macrophages (4), as well as resident memory T cells with both pro- and anti-inflammatory properties (5, 6). The transfer of skin and its resident immune cell populations may underlie the high frequency of rejection episodes in face transplant recipients. At least one episode of skin acute cellular rejection (ACR) has been reported during the first year post-transplant in all face transplant recipients (7), compared to 10 - 15% in kidney or heart recipients (8) on a similar immunosuppressive regimen. Moreover, episodes of ACR after the first year post-transplant are common in face transplants but rare in SOT (9). Although most ACR episodes in VCA are reversible with prompt treatment, reports suggest that the number of ACR episodes may be predictive of future chronic allograft rejection (10) and graft loss (11). A deeper understanding of the immunopathology of face transplant ACR is needed to establish treatment and preventative therapies specifically targeted for these patients. In this study, we addressed this knowledge gap by gene expression profiling, histology, immunostaining and T cell receptor sequencing. Because 5 our primary objective was to investigate the mechanisms of ACR, we studied all grades of ACR that were prospectively diagnosed in one of the most established VCA centers worldwide, thus representing the spectrum of ACR encountered in clinical practice. We also compared rejection with non-transplant related inflammatory skin diseases to identify the genes uniquely associated with face transplant rejection. 6 Results Patient characteristics and biopsy details Between April 2009 and February 2014, seven patients received face transplants at the Brigham and Women’s Hospital (Boston, MA). All patients were transplanted across multiple human leukocyte antigen (HLA) mismatches between donor and recipient (Figure 1A and Supplementary Table 1). There were 5 male and 2 female recipients; all were Caucasian. We identified thirty-eight face transplant skin biopsies from these patients that fulfilled our inclusion criteria (see Methods). These biopsies were examined using multiplex gene expression profiling, immunostaining, histological examination, and T cell receptor sequencing. Histological analysis was performed using the 2007 Banff classification (12) (Grade 0 = no rejection; Grade 1 = mild rejection; Grade 2 = moderate rejection; Grade 3 = severe rejection). All patients had more than one biopsy included with at least one biopsy collected during ACR and one biopsy collected during non-rejection (Figure 1A, B, C and Supplementary Table 2). The mean follow-up time was 76.2 months (Range 48 - 114 months) post-transplant. All episodes of ACR were successfully treated (treatments for each ACR episode are shown in Supplementary Table 2). There were no graft losses or patient deaths. For gene expression analyses, we studied the same archived formalin fixed paraffin embedded (FFPE) skin biopsy used for histologic assessment of rejection; we utilized NanoString based gene expression profiling because of its superior ability to quantitatively measure RNA expression in FFPE samples (13). Severe ACR in face transplants has a distinct gene expression signature We compared the gene expression profiles of Grade 0 biopsies with those obtained during Grade 3 rejection to identify the molecular changes associated with severe ACR. Unsupervised 7 principal component analysis (PCA) clustered Grade 3 biopsies according to similarities in their patterns of expression
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