PMN transendothelial migration decreases nuclear NFκB in IL-1β–activated endothelial cells: role of PECAM-1

During the systemic inflammatory response, circulating cytokines interact with the vascular endothelium, resulting in activation and nuclear accumulation of the nuclear transcription factor, nuclear factor kappa B (NFκB). In turn, NFκB transactivates relevant proinflammatory genes, resulting in an amplification of the inflammatory response. Because this scenario is potentially detrimental to the host, mechanisms exist to limit this amplification. Using an in vitro system that mimics the vascular–interstitial interface during inflammation (cell culture inserts), we provide evidence for the existence of a novel negative feedback mechanism on NFκB activity. We show that the interleukin 1β–induced accumulation of nuclear NFκB in human umbilical vein endothelial cell monolayers is dramatically reduced when polymorphonuclear leukocytes (PMN) are allowed to migrate across these monolayers. This effect does not appear to be due to PMN-derived elastase or nitric oxide. Fixed PMN (adhere but do not migrate) did not affect nuclear NFκB. Furthermore, cross-linking of platelet-endothelial cell adhesion molecule-1 (PECAM-1), but not intercellular adhesion molecule-1, reduces human umbilical vein endothelial cell nuclear NFκB induced by interleukin 1β. Finally, interaction of PMN with PECAM-1–deficient endothelial cells does not reduce nuclear NFκB. These observations indicate that engagement of PECAM-1 by emigrating PMN is a pivotal event in this negative feedback on NFκB activity

Cardiac Fibroblasts Contribute to Myocardial Dysfunction in Mice with Sepsis: The Role of NLRP3 Inflammasome Activation

Myocardial contractile dysfunction in sepsis is associated with the increased morbidity and mortality. Although the underlying mechanisms of the cardiac depression have not been fully elucidated, an exaggerated inflammatory response is believed to be responsible. Nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome is an intracellular platform that is involved in the maturation and release of interleukin (IL)-1 beta. The aim of the present study is to evaluate whether sepsis activates NLRP3 inflammasome/caspase-1/IL-1 beta pathway in cardiac fibroblasts (CFs) and whether this cytokine can subsequently impact the function of cardiomyocytes (cardiac fibroblast-myocyte crosstalk). We show that treatment of CFs with lipopolysaccharide (LPS) induces upregulation of NLRP3, activation of caspase-1, as well as the maturation (activation) and release of IL-1 beta. In addition, the genetic (small interfering ribonucleic acid [siRNA]) and pharmacological (glyburide) inhibition of the NLRP3 inflammasome in CFs can block this signaling pathway. Furthermore, the inhibition of the NLRP3 inflammasome in cardiac fibroblasts ameliorated the ability of LPS-chalenged CFs to impact cardiomyocyte function as assessed by intracellular cyclic adenosine monophosphate (cAMP) responses in cardiomyocytes. Salient features of this the NLP3 inflammasome/ caspase-1 pathway were confirmed in in vivo models of endotoxemia/sepsis. We found that inhibition of the NLRP3 inflammasome attenuated myocardial dysfunction in mice with LPS and increased the survival rate in mice with feces-induced peritonitis. Our results indicate that the activation of the NLRP3 inflammasome in cardiac fibroblasts is pivotal in the induction of myocardial dysfunction in sepsis