Discovery and Validation of a Urinary Exosome mRNA Signature for the Diagnosis of Human Kidney Transplant Rejection.

BACKGROUND: Developing a noninvasive clinical test to accurately diagnose kidney allograft rejection is critical to improve allograft outcomes. Urinary exosomes, tiny vesicles released into the urine that carry parent cells' proteins and nucleic acids, reflect the biologic function of the parent cells within the kidney, including immune cells. Their stability in urine makes them a potentially powerful tool for liquid biopsy and a noninvasive diagnostic biomarker for kidney-transplant rejection. METHODS: Using 192 of 220 urine samples with matched biopsy samples from 175 patients who underwent a clinically indicated kidney-transplant biopsy, we isolated urinary exosomal mRNAs and developed rejection signatures on the basis of differential gene expression. We used crossvalidation to assess the performance of the signatures on multiple data subsets. RESULTS: An exosomal mRNA signature discriminated between biopsy samples from patients with all-cause rejection and those with no rejection, yielding an area under the curve (AUC) of 0.93 (95% CI, 0.87 to 0.98), which is significantly better than the current standard of care (increase in eGFR AUC of 0.57; 95% CI, 0.49 to 0.65). The exosome-based signature's negative predictive value was 93.3% and its positive predictive value was 86.2%. Using the same approach, we identified an additional gene signature that discriminated patients with T cell-mediated rejection from those with antibody-mediated rejection (with an AUC of 0.87; 95% CI, 0.76 to 0.97). This signature's negative predictive value was 90.6% and its positive predictive value was 77.8%. CONCLUSIONS: Our findings show that mRNA signatures derived from urinary exosomes represent a powerful and noninvasive tool to screen for kidney allograft rejection. This finding has the potential to assist clinicians in therapeutic decision making.

T cell-attracting CCL18 chemokine is a dominant rejection signal during limb transplantation.

Limb transplantation is a life-changing procedure for amputees. However, limb recipients have a 6-fold greater rejection rate than solid organ transplant recipients, related in part to greater immunogenicity of the skin. Here, we report a detailed immunological and molecular characterization of individuals who underwent bilateral limb transplantation at our institution. Circulating Th17 cells are increased in limb transplant recipients over time. Molecular characterization of 770 genes in skin biopsies reveals upregulation of T cell effector immune molecules and chemokines, particularly CCL18. Skin antigen-presenting cells primarily express the chemokine CCL18, which binds to the CCR8 receptor. CCL18 treatment recruits more allo-T cells to the skin xenograft in a humanized skin transplantation model, leading to signs of accelerated graft rejection. Blockade of CCR8 remarkedly decreases CCL18-induced allo-T cell infiltration. Our results suggest that targeting the CCL18:CCR8 pathway could be a promising immunosuppressive approach in transplantation.

MMP3 Is a Non-invasive Biomarker of Rejection in Skin-Bearing Vascularized Composite Allotransplantation: A Multicenter Validation Study.

Background: There is unmet need for non-invasive immunomonitoring to improve diagnosis and treatment of acute rejection in vascularized composite allotransplantation (VCA). Circulating matrix metalloproteinase 3 (MMP3) was described as a candidate non-invasive biomarker to predict treatment response to acute rejection in clinical VCA. However, larger validation studies are yet to be reported to allow for more definitive conclusions. Methods: We retrospectively measured MMP3 levels using ELISA in a total of 140 longitudinal serum samples from six internal and three external face transplant recipients, as well as three internal and seven external upper extremity transplant recipients. The control groups comprised serum samples from 36 kidney transplant recipients, 14 healthy controls, and 38 patients with autoimmune skin disease. A linear mixed model was used to study the effect of rejection state (pre-transplant, no-rejection, non-severe rejection (NSR), and severe rejection) on MMP3 levels. Results: In VCA, MMP3 levels increased significantly (p < 0.001) between pre- and post-transplant no-rejection states. A further increase occurred during severe rejection (p < 0.001), while there was no difference in MMP3 levels between non-severe and no-rejection episodes. A threshold of 5-fold increase from pre-transplant levels could discriminate severe from NSR with 76% sensitivity and 81% specificity (AUC = 0.79, 95% CI = 0.65-0.92, p < 0.001). In kidney transplantation, the MMP3 levels were significantly (p < 0.001) elevated during antibody-mediated rejection but not during T-cell mediated rejection (TCMR) (p = 0.547). MMP3 levels in healthy controls and autoimmune skin disease patients were comparable with either pre-transplant or no-rejection/NSR episodes of VCA patients. Conclusion: The results of this study suggest that serum MMP3 protein is a promising marker for stratifying patients according to severity of rejection, complementary to biopsy findings.