Endometriosis is a chronic inflammatory disorder characterized by the presence of endometrial-like tissue growing outside of the uterus. Although the cause is unknown, retrograde menstruation leads to deposition of endometrial cells into the peritoneal cavity. Lack of disease recognition and long diagnostic delays (6-10 years) lead to substantial personal, social and financial burdens, as well as delayed treatment. A non-invasive diagnostic for endometriosis is a major unmet clinical need. Here, we assessed whether differences in menstrual effluent-derived stromal fibroblast cells (ME-SFCs) from women with and without endometriosis provide the basis for a non-invasive diagnostic for endometriosis. In addition, we investigated whether treatment of control ME-SFCs with inflammatory cytokines (TNF and IL-1β) could induce an endometriosis-like phenotype. ME-SFCs from laparoscopically diagnosed endometriosis patients exhibit reduced decidualization capacity, measured by IGFBP1 production after exposure to cAMP. A receiver operating characteristic (ROC) curve developed using decidualization data from controls and endometriosis subjects yielded an area under the curve of 0.92. In addition, a significant reduction in ALDH1A1 gene expression and increased podoplanin surface expression were also observed in endometriosis ME-SFCs when compared to control ME-SFCs. These endometriosis-like phenotypes can be reproduced in control ME-SFCs by exposure to inflammatory cytokines and are associated with increased cell migration. These results are consistent with the hypothesis that chronic intrauterine inflammation influences the development of endometriosis lesions following retrograde menstruation. In conclusion, the analysis of ME-SFCs can provide an accurate, rapid, and non-invasive diagnostic for endometriosis and insight into disease pathogenesis.

Ischemia reperfusion injuries (IRI) are unavoidable in solid organ transplantation. IRI augments alloimmunity but the mechanisms involved are poorly understood. Herein, we examined the effect of IRI on antigen specific alloimmunity. We demonstrate that ischemia promotes alloimmune activation, leading to more severe histological features of rejection, and increased CD4+ and CD8+ T cell graft infiltration, with a predominantly CD8+ IFNγ+ infiltrate. This process is dependent on the presence of alloreactive CD4+ T cells, where depletion prevented infiltration of ischemic grafts by CD8+ IFNγ+ T cells. IL-6 is a known driver of ischemia-induced rejection. Herein, depletion of donor antigen-presenting cells reduced ischemia-induced CD8+ IFNγ+ allograft infiltration, and improved allograft outcomes. Following prolonged ischemia, accelerated rejection was observed despite treatment with CTLA4Ig, indicating that T cell costimulatory blockade failed to overcome the immune activating effect of IRI. However, despite severe ischemic injury, treatment with anti-IL-6 and CTLA4Ig blocked IRI-induced alloimmune injury and markedly improved allograft survival. We describe a novel pathway where IRI activates innate immunity, leading to upregulation of antigen specific alloimmunity, resulting in chronic allograft injury. Based on these findings, we describe a clinically relevant treatment strategy to overcome the deleterious effect of IRI, and provide superior long-term allograft outcomes.

Ischemia reperfusion injury (IRI) evokes intra-graft inflammatory responses, which markedly augment alloimmune responses against the graft. Understanding the mechanisms underlying these responses is fundamental to develop therapeutic regimens to prevent/ameliorate organ IRI. Here, we demonstrate that IRI results in a marked increase in mitochondrial damage and autophagy in dendritic cells (DC). While autophagy is a survival mechanism for ischemic DC, it also augments their production of IL6. Allograft derived dendritic cells (ADDC) lacking autophagy related gene 5 (Atg5) showed higher death rates post-transplantation. Transplanted ischemic hearts from CD11cCre/Atg5 conditional knockout mice showed marked reduction in intra-graft expression of IL6 as compared to controls. To antagonize the effect of IL6 locally in the heart, we synthesized novel anti-IL6 nanoparticles with capacity for controlled release of anti-IL6 over time. As compared to systemic delivery of anti-IL6, localized delivery of anti-IL6 significantly reduced chronic rejection with a markedly lower amount administered. Despite improved allograft histology, there were no changes to splenic T cell populations, illustrating the importance of local IL6 in driving chronic rejection after IRI. These data carry potential clinical significance, by identifying an innovative, targeted strategy to manipulate organs prior to transplantation to diminish inflammation, leading to improved long term outcomes.