Comunicación

DANGER SIGNALS IN THE ORGAN PRESERVATION SOLUTION ACTIVATE MYELOID NLRP3 INFLAMMASOME: IMPACT OF COLD ISCHEMIA TIME AND TYPE OF ORGAN DONATION AND INFLUENCING ON THE SHORT-TERM OUTCOME OF LIVER TRANSPLANTATION.

Autores:

FERNANDO LUCAS RUIZ1, Francisco Villalba López2, CARLOS DE TORRE MINGUELA2, Felipe Alconchel Gago2, MARTA JOVER AGUILAR3, Laura Martinez Alarcon2, Maria Isabel Sanchez Lorencio4, Antonio Rios Zambudio2, Pablo Ramirez Romero2, JOSE ANTONIO PONS MIÑANO2, PABLO PELEGRIN VIVANCOS2, ALBERTO BAROJA MAZO2

Afiliaciones:

(1) OFTALMOLOGÍA EXPERIMENTAL, IMIB-Arrixaca, España
(2) CIRUGÍA DIGESTIVA, ENDOCRINA Y TRASPLANTE DE ÓRGANOS ABDOMINALES, IMIB-Arrixaca, España
(3) Servicio de Cirugía. Unidad de Trasplantes. Hospital Clínico Universitario Virgen de la Arrixaca, 30120, España (Región de Murcia)
(4) Inflammation Research Group, España (Región de Murcia)

Comunicación:

Antecedentes:

The innate system is prepared to sense "danger signals", described as DAMPs (Danger-associated molecular patterns) and respond to them, generally by creating a pro-inflammatory environment. These DAMPs would be formed mainly by self-molecules of the organism that are not usually found extracellularly and that would appear after tissue damage. Although by different pathways, all these DAMPs are able to activate the formation of the inflammasome in monocytes/macrophages. The inflammasome is a multiprotein complex formed by a sensor, being NLR Family Pyrin Domain Containing 3 (NLRP3) the most significant, which recruits PYD and CARD Domain Containing (ASC), which in turn recruits pro-caspase 1. Caspase 1 (Casp1) processes and matures interleukin (IL)-1beta which acts in an autocrine and paracrine manner on the tissue, causing an inflammatory environment, which can influence the rejection machinery. Here we show how the type of donation (after brain death or after circulatory death), or even the cold ischemia time (CIT), had a high incidence in the presence of DAMPs in the organ preservation solution (OPS) and how these DAMPs were able to activate the inflammasome in myeloid cells.

Métodos:

We have quantified the presence of several DAMPs in the OPS after ischemia, prior to revascularization, such as High Mobility Group Box 1 (HMGB1) alarmine, hyaluronan or fibronectin among others, in addition to pro-inflammatory cytokines. Both inflammasome formation and activation in human primary monocytes and human monocyte THP-1 cell line were analysed by ELISA, WB, PCR and immunolabelling.

Resultados:

We have verified how OPS is capable of activating the release of IL-1β in THP1 cells. This release was significantly reduced when we inhibited pharmacologically NLRP3 or Casp1 or when we used THP1 Knock-Out cells for the NLRP3, ASC or Casp1 inflammasome components. We also found that THP1 cells, after incubation with OPS, assemble the inflammatory complex in the form of specks (Figure 1). Likewise, primary human monocytes activated the inflammasome when incubated with OPS. Nevertheless, OPS was able to prime monocytes through NF-κB pathway. Large number of DAMPs were detected in OPS after cold ischemia (Figure 2). HMGB1, uric acid, fibronectin, HSP70, ASC specks and endotoxins had a positive correlation with the release of IL-1β from THP-1 cells after OPS incubation. Several detected DAMPS had a great correlation with cold ischemia time, including HSP-70, IL-18, or ASC specks, as well as active caspases 1, 3 and 8 and even the damage marker LDH. Likewise, ASC specks, HSP70 and endotoxins were differentially released to OPS after brain death or circulatory death. Interestingly, we also found a relationship between the prognosis of the transplants and the DAMPs found in their own OPS.

Conclusiones:

Altogether, these data indicate that OPS after ischemia has a high amount of potential DAMPs, which may mediate activation of NLRP3 inflammasome in monocytes. Furthermore, there is a correlation between the presence of these DAMPs in the OPS and possible liver rejection. Therefore, the analysis and quantification of DAMPs in the post-ischemic graft could provide a predictive value of the progression of the transplant in the short time. Moreover, new inhibitors directed against inflammasome pathway could be developed to ameliorate the damage produced by ischemia in donated organs.


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Dirección

Campus de Ciencias de la Salud
Carretera Buenavista s/n, 30120 El Palmar
Murcia, España

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