Subcellular compartmentalization of reactive oxygen species (ROS) plays a critical role in transmitting cell signals in response to environmental stimuli. In this regard, signals at the plasma membrane have been shown to trigger NADPH oxidase-dependent ROS production within the endosomal compartment and this step can be required for redox-dependent signal transduction. Unique features of redox-active signaling endosomes can include NADPH oxidase complex components (Nox1, Noxo1, Noxa1, Nox2, p47phox, p67phox, and=or Rac1), ROS processing enzymes (SOD1 and=or peroxiredoxins), chloride channels capable of mediating superoxide transport and=or membrane gradients required for Nox activity, and novel redox-dependent sensors that control Nox activity. This review will discuss the cytokine and growth factor receptors that likely mediate signaling through redox-active endosomes, and the common mechanisms whereby they act. Additionally, the review will cover ligand-independent environmental injuries, such as hypoxia=reoxygenation injury, that also appear to facilitate cell signaling through NADPH oxidase at the level of the endosome. We suggest that redox-active endosomes encompass a subset of signaling endosomes that we have termed redoxosomes. Redoxosomes are uniquely equipped with redox-processing proteins capable of transmitting ROS signals from the endosome interior to redox-sensitive effectors on the endosomal surface. In this manner, redoxosomes can control redoxdependent effector functions through the spatial and temporal regulation of ROS as second messengers. Antioxid. Redox Signal. 11, 1313-1333

Growing evidence suggests that NADPH oxidase (Nox)-derived reactive oxygen species (ROS) play important roles in regulating cytokine signaling. We have explored how TNF-a induction of Nox-dependent ROS influences NF-kB activation. Cellular stimulation by TNF-a induced NADPH-dependent superoxide production in the endosomal compartment, and this ROS was required for IKK-mediated activation of NF-kB. Inhibiting endocytosis reduced the ability of TNF-a to induce both NADPH-dependent endosomal superoxide and NF-kB, supporting the notion that redox-dependent signaling of the receptor occurs in the endosome. Molecular analyses demonstrated that endosomal H 2 O 2 was critical for the recruitment of TRAF2 to the TNFR1=TRADD complex after endocytosis. Studies using both Nox2 siRNA and Nox2-knockout primary fibroblasts indicated that Nox2 was critical for TNF-a-mediated induction of endosomal superoxide. Redox-active endosomes that form after TNF-a or IL-1b induction recruit several common proteins (Rac1, Nox2, p67 phox , SOD1), while also retaining specificity for ligand-activated receptor effectors. Our studies suggest that TNF-a and IL-1b signaling pathways both can use Nox2 to facilitate redox activation of their respective receptors at the endosomal level by promoting the redox-dependent recruitment of TRAFs. These studies help to explain how cellular compartmentalization of redox signals can be used to direct receptor activation from the plasma membrane.

To our knowledge this is the first study to provide detailed comparisons of acute transfusion reactions to all blood products between pediatric and adult populations at a single institution and supported by a single transfusion service and culture. Collectively these data provide insight into pediatric transfusion reactions and demonstrate a general increase in the incidence of transfusion reactions within the pediatric compared to adult population.

Recent evidence suggests that signaling by the proinflammatory cytokine interleukin-1␤ (IL-1␤) is dependent on reactive oxygen species derived from NADPH oxidase. Redox signaling in response to IL-1␤ is known to require endocytosis of its cognate receptor (IL-1R1) following ligand binding and the formation of redox-active signaling endosomes that contain Nox2 (also called redoxosomes). The consequent generation of reactive oxygen species by redoxosomes is responsible for the downstream recruitment of IL-1R1 effectors (IRAK, TRAF6, and IB kinase kinases) and ultimately for activation of the transcription factor NFB. Despite this knowledge of the signaling events that occur downstream of redoxosome formation, an understanding of the mechanisms that coordinate the genesis of redoxosomes following IL-1␤ stimulation has been lacking. Here, we demonstrate that lipid rafts play an important role in this process. We show that Nox2 and IL-1R1 localize to plasma membrane lipid rafts in the unstimulated state and that IL-1␤ signals caveolin-1-dependent endocytosis of both proteins into the redoxosome. We also show that inhibiting lipid raft-mediated endocytosis prevents NFB activation. Finally, we demonstrate that Vav1, a Rac1 guanine exchange factor and activator of Nox2, is recruited to lipid rafts following IL-1␤ stimulation and that it is required for NFB activation. Our results fill in an important mechanistic gap in the understanding of early IL-1R1 and Nox2 signaling events that control NFB activation, a redox-dependent process important in inflammation. IL-1␤2 is a potent proinflammatory cytokine that controls inflammation in response to a diverse collection of health problems, including ischemia/reperfusion injury, viral infection, bacterial infection, autoimmune diseases such as diabetes, allergies, trauma, and chemical exposure (1-8). A primary role for signaling by IL-1␤ in these inflammatory responses is the activation of NFB, a transcription factor that regulates a large number of immune molecules, apoptotic factors, anti-apoptotic factors, and other transcription factors (9). The importance of IL-1␤ and NFB in inflammation was highlighted by a clinical study on mortality in septic patients (10). A spectrum of cytokines and transcription factors was examined in this study, and two were identified as significant prognostic indictors of patient outcome. One prognostic indicator was NFB activity, with this transcription factor twice as active in non-survivors relative to survivors. The second prognostic indicator kinase was the ratio between IL-1␤ and its competitive antagonist IL-1ra, with survivors having a 50% higher IL-1ra/ IL-1␤ ratio than non-survivors.Because of the clinical importance of IL-1␤, elucidating the signaling events involved in IL-1␤-mediated NFB activation is of great significance. Among the early events that control IL-1␤ signaling is the induction of IL-1R1 dimerization following ligand binding (11,12). This event initiates binding of MyD88 to the TIR (Toll/IL-1R1) domains within the cytopl...

The AP-1 transcription factor modulates a wide range of cellular processes, including cellular proliferation, programmed cell death, and survival. JunD is a major component of the AP-1 complex following liver ischemia/reperfusion (I/R) injury; however, its precise function in this setting remains unclear. We investigated the functional significance of JunD in regulating AP-1 transcription following partial lobar I/R injury to the liver, as well as the downstream consequences for hepatocellular remodeling. Our findings demonstrate that JunD plays a protective role, reducing I/R injury to the liver by suppressing acute transcriptional activation of AP-1. In the absence of JunD, c-Jun phosphorylation and AP-1 activation in response to I/R injury were elevated, and this correlated with increased caspase activation, injury, and alterations in hepatocyte proliferation. The expression of dominant negative JNK1 inhibited c-Jun phosphorylation, AP-1 activation, and hepatic injury following I/R in JunD ؊/؊ mice but, paradoxically, led to an enhancement of AP-1 activation and liver injury in JunD ؉/؊ littermates. Enhanced JunD/JNK1-dependent liver injury correlated with the acute induction of diphenylene iodonium-sensitive NADPHdependent superoxide production by the liver following I/R. In this context, dominant negative JNK1 expression elevated both Nox2 and Nox4 mRNA levels in the liver in a JunD-dependent manner. These findings suggest that JunD counterbalances JNK1 activation and the downstream redox-dependent hepatic injury that results from I/R, and may do so by regulating NADPH oxidases.