Reduced CXCL1 production by endogenous IL-37 expressing dendritic cells does not affect T cell activation.

The dendritic cell (DC)-derived cytokine profile contributes to naive T cell differentiation, thereby directing the immune response. IL-37 is a cytokine with anti-inflammatory characteristics that has been demonstrated to induce tolerogenic properties in DC. In this study we aimed to evaluate the influence of IL-37 on DC-T cell interaction, with a special focus on the role of the chemokine CXCL1. DC were cultured from bone marrow of human IL-37 transgenic (hIL-37Tg) or WT mice. The phenotype of unstimulated and LPS-stimulated DC was analyzed (co-stimulatory molecules and MHCII by flow cytometry, cytokine profile by RT-PCR and ELISA), and T cell stimulatory capacity was assessed in mixed lymphocyte reaction. The role of CXCL1 in T cell activation was analyzed in T cell stimulation assays with anti-CD3 or allogeneic DC. The expression of the co-stimulatory molecules CD40, CD80 and CD86, and of MHCII in LPS-stimulated DC was not affected by endogenous expression of IL-37, whereas LPS-stimulated hIL-37Tg DC produced less CXCL1 compared to LPS-stimulated WT DC. T cell stimulatory capacity of LPS-matured hIL-37Tg DC was comparable to that of WT DC. Recombinant mouse CXCL1 did not increase T cell proliferation either alone or in combination with anti-CD3 or allogeneic DC, nor did CXCL1 affect the T cell production of interferon-γ and IL-17. Endogenous IL-37 expression does not affect mouse DC phenotype or subsequent T cell stimulatory capacity, despite a reduced CXCL1 production. In addition, we did not observe an effect of CXCL1 in T cell proliferation or differentiation

Ablation of NLRP3, but not ASC, abrogated IPC protective effects on cardiac function following I/R.

<p>Effect of ASC and NLRP3 gene ablation on cardiac mechanical performance following acute I/R with or without preceding IPC in isolated perfused mouse hearts. (A) End-diastolic pressure (EDP) measured at end reperfusion for the different groups; (B) % Rate-pressure product (RPP) determined at end reperfusion and normalized to baseline, pre-ischemic, values for the different groups. Black bars denote I/R groups, white bars reflect IPC + I/R groups. (n = 7 for all groups). Mean ± SEM, * P<0.05 vs. I/R in WT, ^# P<0.05 IPC+I/R vs. I/R similar group (ANOVA).</p

Unaltered HMGB1 mRNA levels.

<p>Myocardial HMGB1 mRNA levels at end reperfusion for wild-type (WT), ASC^−/− and NLRP3^−/− hearts subjected to ischemia-reperfusion with or without IPC. Black bars denote I/R groups, white bars reflect IPC + I/R groups. (n = 7 for all groups). Mean ± SEM, * P<0.05 vs. I/R in WT (ANOVA).</p

Decreased STAT3 and increased AMPK protein levels in NLRP3^−/− hearts.

<p>Effect of NLRP3 gene ablation on survival kinases and their phosphorylation status in preconditioned hearts analyzed at 5 min reperfusion following 35 min ischemia. PKC-ε (A), ERK (C), AMPK (E) and STAT3 (G), and their phosphorylation status (B, D, F, and H, respectively) are shown (n = 4 hearts per genotype). Mean ± SEM, *P<0.05 WT vs. NLRP3^−/−.</p

Reduced IL-6 cytokine levels in reperfused NLRP3^−/− hearts.

<p>Myocardial cytokine levels at end reperfusion for wild-type (WT), ASC^−/− and NLRP3^−/− hearts subjected to I/R with or without IPC. IL-6 (A), TNF-α (B) and IL-1β (C) are given. Black bars denote I/R groups, white bars reflect IPC + I/R groups. (n = 7 for all groups). Mean ± SEM, * P<0.05 vs. I/R in WT (ANOVA).</p

Physiological characteristics of Langendorff-perfused hearts of wild-type (WT), ASC^−/− and NLRP3^−/− mice.

<p>Values are given as (mean ± SEM). EDP: end-diastolic pressure; Psys: peak systolic pressure; HR: heart rate; RPP: rate-pressure-product.</p

Deletion of the innate immune NLRP3 receptor abolishes cardiac ischemic preconditioning and is associated with decreased Il-6/STAT3 signaling

Contains fulltext : 110810.pdf (publisher's version ) (Open Access)OBJECTIVE: Recent studies indicate that the innate immune system is not only triggered by exogenous pathogens and pollutants, but also by endogenous danger signals released during ischemia and necrosis. As triggers for the innate immune NLRP3 inflammasome protein complex appear to overlap with those for cardiac ischemia-reperfusion (I/R) and ischemic preconditioning (IPC), we explored the possibility that the NLRP3 inflammasome is involved in IPC and acute I/R injury of the heart. PRINCIPAL FINDINGS: Baseline cardiac performance and acute I/R injury were investigated in isolated, Langendorff-perfused hearts from wild-type (WT), ASC(-/-) and NLRP3(-/-) mice. Deletion of NLRP3 inflammasome components ASC(-/-) or NLRP3(-/-) did not affect baseline performance. The deletions exacerbated I/R-induced mechanical dysfunction, but were without effect on I/R-induced cell death. When subjected to IPC, WT and ASC(-/-) hearts were protected against I/R injury (improved function and less cell death). However, IPC did not protect NLRP3(-/-) hearts against I/R injury. NLRP3(-/-) hearts had significantly decreased cardiac IL-6 levels with a trend towards lower IL-1beta levels at end reperfusion, suggesting abrogation of IPC through diminished IL-6 and/or IL-1beta signaling. Subsequent experiments showed that neutralising IL-6 using an antibody against IL-6 abrogated IPC in WT hearts. However, inhibition of the IL-1r receptor with the IL-1 receptor inhibitor Anakinra (100 mg/L) did not abrogate IPC in WT hearts. Analysis of survival kinases after IPC demonstrated decreased STAT3 expression in NLRP3(-/-) hearts when compared to WT hearts. CONCLUSIONS: The data suggest that the innate immune NLRP3 protein, in an NLRP3-inflammasome-independent fashion, is an integral component of IPC in the isolated heart, possibly through an IL-6/STAT3 dependent mechanism