Intracellular lipopolysaccharide from Gram-negative bacteria including Escherichia coli, Salmonella typhimurium, Shigella flexneri, and Burkholderia thailandensis activates mouse caspase-11, causing pyroptotic cell death, interleukin-1β processing, and lethal septic shock. How caspase-11 executes these downstream signalling events is largely unknown. Here we show that gasdermin D is essential for caspase-11-dependent pyroptosis and interleukin-1β maturation. A forward genetic screen with ethyl-N-nitrosourea-mutagenized mice links Gsdmd to the intracellular lipopolysaccharide response. Macrophages from Gsdmd(-/-) mice generated by gene targeting also exhibit defective pyroptosis and interleukin-1β secretion induced by cytoplasmic lipopolysaccharide or Gram-negative bacteria. In addition, Gsdmd(-/-) mice are protected from a lethal dose of lipopolysaccharide. Mechanistically, caspase-11 cleaves gasdermin D, and the resulting amino-terminal fragment promotes both pyroptosis and NLRP3-dependent activation of caspase-1 in a cell-intrinsic manner. Our data identify gasdermin D as a critical target of caspase-11 and a key mediator of the host response against Gram-negative bacteria.

Phenotypic modulation of endothelium to a dysfunctional state contributes to the pathogenesis of cardiovascular diseases such as atherosclerosis. The localization of atherosclerotic lesions to arterial geometries associated with disturbed flow patterns suggests an important role for local hemodynamic forces in atherogenesis. There is increasing evidence that the vascular endothelium, which is directly exposed to various fluid mechanical forces generated by pulsatile blood flow, can discriminate among these stimuli and transduce them into genetic regulatory events. At the level of individual genes, this regulation is accomplished via the binding of certain transcription factors, such as NFκB and Egr‐1, to shear‐stress response elements (SSREs) that are present in the promoters of biomechanically inducible genes. At the level of multiple genes, distinct patterns of up‐ and downregulation appear to be elicited by exposure to steady laminar shear stresses versus comparable levels of non‐laminar (e.g., turbulent) shear stresses or cytokine stimulation (e.g., IL‐1β). Certain genes upregulated by steady laminar shear stress stimulation (such as eNOS, COX‐2, and Mn‐SOD) support vasoprotective (i.e., anti‐inflammatory, anti‐thrombotic, anti‐oxidant) functions in the endothelium. We hypothesize that the selective and sustained expression of these and related “atheroprotective genes” in the endothelial lining of lesion‐protected areas represents a mechanism whereby hemodynamic forces can influence lesion formation and progression.

One of the striking features of vascular endothelium, the single-cellthick lining of the cardiovascular system, is its phenotypic plasticity. Various pathophysiologic factors, such as cytokines, growth factors, hormones, and metabolic products, can modulate its functional phenotype in health and disease. In addition to these humoral stimuli, endothelial cells respond to their biomechanical environment, although the functional implications of this biomechanical paradigm of activation have not been fully explored. Here we describe a highthroughput genomic analysis of modulation of gene expression observed in cultured human endothelial cells exposed to two well defined biomechanical stimuli-a steady laminar shear stress and a turbulent shear stress of equivalent spatial and temporal average intensity. Comparison of the transcriptional activity of 11,397 unique genes revealed distinctive patterns of up-and down-regulation associated with each type of stimulus. Cluster analyses of transcriptional profiling data were coupled with other molecular and cell biological techniques to examine whether these global patterns of biomechanical activation are translated into distinct functional phenotypes. Confocal immunofluorescence microscopy of structural and contractile proteins revealed the formation of a complex apical cytoskeleton in response to laminar shear stress. Cell cycle analysis documented different effects of laminar and turbulent shear stresses on cell proliferation. Thus, endothelial cells have the capacity to discriminate among specific biomechanical forces and to translate these input stimuli into distinctive phenotypes. The demonstration that hemodynamically derived stimuli can be strong modulators of endothelial gene expression has important implications for our understanding of the mechanisms of vascular homeostasis and atherogenesis.