Characterization of Transfusion-Elicited Acute Antibody-Mediated Rejection in a Rat Model of Kidney Transplantation

Abstract

Animal models of antibody-mediated rejection (ABMR) may provide important evidence supporting proof of concept. We elicited donor-specific antibodies (DSA) by transfusion of donor blood (Brown Norway RT1n) into a complete mismatch recipient (Lewis RT1l) 3 weeks prior to kidney transplantation. Sensitized recipients had increased anti-donor splenocyte IgG1, IgG2b and IgG2c DSA 1 week after transplantation. Histopathology was consistent with ABMR characterized by diffuse peritubular capillary C4d and moderate microvascular inflammation with peritubular capillaritis + glomerulitis > 2. Immunofluorescence studies of kidney allograft tissue demonstrated a greater CD68/CD3 ratio in sensitized animals, primarily of the M1 (pro-inflammatory) phenotype, consistent with cytokine gene analyses that demonstrated a predominant T helper (TH)1 (interferon-γ, IL-2) profile. Immunoblot analyses confirmed the activation of the M1 macrophage phenotype as interferon regulatory factor 5, inducible nitric oxide synthase and phagocytic NADPH oxidase 2 were significantly up-regulated. Clinical biopsy samples in sensitized patients with acute ABMR confirmed the dominance of M1 macrophage phenotype in humans. Despite the absence of tubulitis, we were unable to exclude the effects of T cell–mediated rejection. These studies suggest that M1 macrophages and TH1 cytokines play an important role in the pathogenesis of acute mixed rejection in sensitized allograft recipients.

Triptolide inhibits donor-specific antibody production and attenuates mixed antibody-mediated renal allograft injury

“…In addition to B cells, other inflammatory cells such as macrophages, T cells, and NK cells are also well known to infiltrate renal allografts during AMR and have proved to jointly contribute to AMR allograft injuries . In accordance with other studies and our previous results showing that macrophages are the most predominant inflammatory cells infiltrating the grafts of AMR, and that M1 macrophages play a dominant role in the pathogenesis of AMR, we demonstrated here that graft‐infiltrating macrophages predominantly polarized to M1 subsets in acute AMR, whereas the absolute numbers of total macrophages as well as M1 and M2 cells in grafts were dramatically reduced when treated with triptolide, and the rise of M2/Mφ ratio may be attributed to the stronger suppressive effect of triptolide on M1. Given that IFN‐γ and IL‐4 are of great importance in driving Mφ to M1 and M2 polarization and enhancing their activities, triptolide remarkably reduced mRNA expression levels of both IFN‐γ and IL‐4, implying that triptolide inhibits macrophage‐mediated responses by synergistically reducing their numbers as well as their functional enhancers.…”

Precision Sub-phenotyping of Histologically Stable Kidney Transplants by a Composite Molecular and Cellular Instability Score

“…Increased numbers of M2 than M1 macrophages have been found in some studies to correlate with graft-loss [99]. KLF4 expression in increased in the InstaScore, and interestingly this gene plays a key role to polarize macrophage into the M2 subset [100], which undertake host defense and wound healing/tissue remodeling tasks [101] and thus can be found to be enriched in renal fibrosis [102].In the AR bopsies in this study, as expected, M1 macrophage signatures were higher in AR [103]. Thus, the macrophage subsets show a dynamic state of skewing the preponderance of M2 to M1 macrophages in the process of immunological priming from pre-AR and full blown AR, and the InstaScore uncovers the state of AR priming or pre-AR in histologically and clinically stable kidney transplants, further highlighting the value of identifying hSTA grafts that are thus at greater risk of AR over time.…”

Markers of Endothelial-to-Mesenchymal Transition

“…An experimental model of ABMR was developed in rats, where DSA were elicited by transfusion of donor blood (Brown Norway RT1n) into a complete mismatch recipient (Lewis RT1l) 3 weeks prior to kidney transplantation, 29 and the renal grafts were studied 7 days posttransplant. In the present study, paraffin-embedded sections of control or ABMR kidneys were used to detect the expression of EndMT markers in PTC.…”