Granzyme A inhibition reduces inflammation and increases survival during abdominal sepsis.

Aims: Peritonitis is one of the most common causes of sepsis, a serious syndrome characterized by a dysregulated systemic inflammatory response. Recent evidence suggests that Granzyme A (GzmA), a serine protease mainly expressed by NK and T cells, could act as a proinflammatory mediator and could play an important role in the pathogenesis of sepsis. This work aims to analyze the role and the therapeutic potential of GzmA in the pathogenesis of peritoneal sepsis. Methods: The level of extracellular GzmA as well as GzmA activity were analyzed in serum from healthy volunteers and patients with confirmed peritonitis and were correlated with the Sequential Organ Failure Assessment (SOFA) score. Peritonitis was induced in C57Bl/6 (WT) and GzmA-/- mice by cecal ligation and puncture (CLP). Mice were treated intraperitoneally with antibiotics alone or in combination serpinb6b, a specific GzmA inhibitor, for 5 days. Mouse survival was monitored during 14 days, levels of some proinflammatory cytokines were measured in serum and bacterial load and diversity was analyzed in blood and spleen at different times. Results: Clinically, elevated GzmA was observed in serum from patients with abdominal sepsis suggesting that GzmA plays an important role in this pathology. In the CLP model GzmA deficient mice, or WT mice treated with an extracellular GzmA inhibitor, showed increased survival, which correlated with a reduction in proinflammatory markers in both serum and peritoneal lavage fluid. GzmA deficiency did not influence bacterial load in blood and spleen and GzmA did not affect bacterial replication in macrophages in vitro, indicating that GzmA has no role in bacterial control. Analysis of GzmA in lymphoid cells following CLP showed that it was mainly expressed by NK cells. Mechanistically, we found that extracellular active GzmA acts as a proinflammatory mediator in macrophages by inducing the TLR4-dependent expression of IL-6 and TNFα. Conclusions: Our findings implicate GzmA as a key regulator of the inflammatory response during abdominal sepsis and provide solid evidences about its therapeutic potential for the treatment of this severe pathology.

The Multifaceted Function of Granzymes in Sepsis: Some Facts and a Lot to Discover.

Sepsis is a serious global health problem. In addition to a high incidence, this syndrome has a high mortality and is responsible for huge health expenditure. The pathophysiology of sepsis is very complex and it is not well-understood yet. However, it is widely accepted that the initial phase of sepsis is characterized by a hyperinflammatory response while the late phase is characterized by immunosuppression and immune anergy, increasing the risk of secondary infections. Granzymes (Gzms) are a family of serine proteases classified according to their cleavage specificity. Traditionally, it was assumed that all Gzms acted as cytotoxic proteases. However, recent evidence suggests that GzmB is the one with the greatest cytotoxic capacity, while the cytotoxicity of others such as GzmA and GzmK is not clear. Recent studies have found that GzmA, GzmB, GzmK, and GzmM act as pro-inflammatory mediators. Specially, solid evidences show that GzmA and GzmK function as extracellular proteases that regulate the inflammatory response irrespectively of its ability to induce cell death. Indeed, studies in animal models indicate that GzmA is involved in the cytokine release syndrome characteristic of sepsis. Moreover, the GZM family also could regulate other biological processes involved in sepsis pathophysiology like the coagulation cascade, platelet function, endothelial barrier permeability, and, in addition, could be involved in the immunosuppressive stage of sepsis. In this review, we provide a comprehensive overview on the contribution of these novel functions of Gzms to sepsis and the new therapeutic opportunities emerging from targeting these proteases for the treatment of this serious health problem.

Intensive Care to Facilitate Organ Donation: A Report on the Experience of 2 Spanish Centers With a Common Protocol.

BACKGROUND: The aim of this study is to report the experience with a program of Intensive Care to facilitate Organ Donation (ICOD) in 2 Spanish centers based on a common protocol. METHODS: Retrospective review of clinical charts of patients with a devastating brain injury whose families were approached to discuss the possibility of ICOD once further treatment was deemed futile by the treating team. Study period is from January 1, 2011, to December 31, 2015. RESULTS: ICOD was discussed with families of 131 patients. Mean age of possible donors was 75 years (SD = 11 years). The main cause of brain injury was an intracranial hemorrhage (72%). Interviews with families were held after the decision had been made not to intubate/ventilate in 50% of cases, and after the decision not to continue with invasive ventilation in the remaining cases. Most interviews (66%) took place in the emergency department. The majority of families (95%) consented to ICOD. Of the 125 consented cases, 101 (81%) developed brain death (BD), most in 72 hours or less. Ninety-nine (98%) patients transitioned to actual donation after BD, with 1.2 organs transplanted per donor. Of patients who did not evolve to BD, 4 died after an unexpected cardiac arrest and 18 after the withdrawal of life-sustaining measures. ICOD contributed to 33% of actual donors registered at both centers. CONCLUSIONS: ICOD is well accepted by families. Most patients evolve to BD within a short period of time. The practice substantially contributes to increasing organ donation and offers more patients the chance of donating their organs after death.