Systemic inflammation in xenograft recipients precedes activation of coagulation

Abstract

Background Dysregulation of coagulation is considered a major barrier against successful pig organ xenotransplantation in nonhuman primates. Inflammation is known to promote activation of coagulation. The role of pro-inflammatory factors as well as the relationship between inflammation and activation of coagulation in xenograft recipients is poorly understood. Methods Baboons received kidney (n=3), heart (n=4) or artery patch (n=8) xenografts from α1,3-galactosyltransferase gene-knockout (GTKO) pigs or GTKO pigs additionally transgenic for human complement regulatory protein CD46 (GTKO/CD46). Immunosuppression (IS) was based on either CTLA4-Ig or anti-CD154 costimulation blockade. Three artery patch recipients did not receive IS. Pro-inflammatory cytokines, chemokines and coagulation parameters were evaluated in the circulation after transplantation. In artery patch recipients, monocytes and dendritic cells (DC) were monitored in peripheral blood. Expression of tissue factor (TF) and CD40 on monocytes and DC were assessed by flow cytometry. C-reactive protein (C-RP) levels in the blood and C-RP deposition in xenografts as well as native organs were evaluated. Baboon and pig C-RP mRNA in heart and kidney xenografts were evaluated. Results In heart and kidney xenograft recipients, the levels of INFγ, TNF-α, IL-12 and IL-8 were not significantly higher after transplantation. However, MCP-1 and IL-6 levels were significantly higher after transplantation, particularly in kidney recipients. Elevated C-RP levels preceded activation of coagulation in heart and kidney recipients, where high levels of C-RP were maintained until the time of euthanasia in both heart and kidney recipients. In artery patch recipients, INFγ, TNF-α, IL-12, IL-8 and MCP-1 were elevated with no IS, while IL-6 was not. With IS, INFγ, TNF-α, IL-12, IL-8 and MCP-1 were reduced, but IL-6 was elevated. Elevated IL-6 levels were observed as early as 2 weeks in artery patch recipients. While IS was associated with reduced thrombin activation, fibrinogen and C-RP levels were increased when IS was given. There was a significant positive-correlation between C-RP, IL-6, and fibrinogen levels. Additionally, absolute numbers of monocytes were significantly increased when IS was given, but not without IS. This was associated with increased CD40 and TF expression on CD14+ monocytes and lineageneg CD11c+ DC, with increased differentiation of the pro-inflammatory CD14+ CD11c+ monocyte population. At the time of euthanasia, C-RP deposition in kidney and heart xenografts, C-RP positive cells in artery patch xenograft and native lungs were detected. Finally, high levels of both pig and baboon C-RP mRNA were detected in heart and kidney xenografts. Conclusions Inflammatory responses precede activation of coagulation after organ xenotransplantation. Early upregulation of C-RP and IL-6 levels may amplify activation of coagulation through upregulation of TF on innate immune cells. Prevention of systemic inflammation in xenograft recipients (SIXR) may be required ...

Silencing Porcine CMAH and GGTA1 Genes Significantly Reduces Xenogeneic Consumption of Human Platelets by Porcine Livers

“…12-15 Recent efforts to reduce xenogeneic thrombocytopenia in a pig-to-primate model have also exposed the importance of limiting immune-mediated graft injury. 16,17 Judicious immunosuppression has prevented thrombocytopenia following pig-to-primate xenotransplantation of the porcine heart and kidney. 18,19 Given this information, it is possible that a combination of immunosuppression and the reduced antigenicity of newer knockout porcine donors alone may facilitate success for renal or cardiac xenografts.…”

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“…A prolonged and persistent inflammatory response to even a small pig xenograft, for example, an artery patch, was demonstrated to precede the coagulation disturbance [21]. As inflammation can augment an immune response, the inflammatory response has been suppressed by the administration of agents such as the interleukin-6 receptor blocker, tocilizumab, and the anti-tumor necrosis factor-alpha agent, etanercept.…”

Synthetic Biology in Cell and Organ Transplantation

“…Activation of the coagulation cascade is a consequence of the inflammatory responses described above, leading to disseminated intravascular coagulation and thrombosis, rapid loss of platelet and clotting factors, decreased blood flow and ischemia/infarction (Schmelzle et al 2010;Ezzelarab et al 2015b). As with inflammation, coagulation is rapidly amplified after initiation and difficult to regulate once widespread.…”