Dendritic cells (DCs) are the most potent APCs. Whereas immature DCs downregulate T-cell responses to induce/ maintain immunologic tolerance, mature DCs promote immunity. To amplify their functions, DCs communicate with neighboring DCs through soluble mediators, cell-to-cell contact, and vesicle exchange. Transfer of nanovesicles (< 100 nm) derived from the endocytic pathway (termed exosomes) represents a novel mechanism of DC-to-DC communication. The facts that exosomes contain exosomeshuttle miRNAs and DC functions can be regulated by exogenous miRNAs, suggest that DC-to-DC interactions could be mediated through exosome-shuttle miRNAs, a hypothesis that remains to be tested. Importantly, the mechanism of transfer of exosome-shuttle miRNAs from the exosome lumen to the cytosol of target cells is unknown. Here, we demonstrate that DCs release exosomes with different miRNAs depending on the maturation of the DCs. By visualizing spontaneous transfer of exosomes between DCs, we demonstrate that exosomes fused with the target DCs, the latter followed by release of the exosome content into the DC cytosol. Importantly, exosome-shuttle miRNAs are functional, because they repress target mRNAs of acceptor DCs. IntroductionCellular miRNAs are released membrane free 1 or packaged inside microvesicles (0.1-1 m) shed by the plasma membrane 2,3 or within nanovesicles (Ͻ 100nm) derived from the endocytic pathway known as exosomes. 4,5 Exosomes are generated as intraluminal vesicles by reverse budding of the membrane of multivesicular bodies (MVBs). Release of exosomes occurs when MVBs fuse their limiting membrane with the plasma membrane. [6][7][8][9] Dendritic cells (DCs) are APCs with the ability to regulate adaptive immunity. Whereas immature DCs down-regulate T-cell responses, mature DCs promote activation, proliferation, and differentiation of effector T cells. 10 Communication between DCs is essential to amplify their tolerogenic and immunogenic functions. 11,12 This DC-to-DC interaction is mediated through cell-tocell contact, soluble mediators, exchange of plasma membrane patches, 13,14 nanotubules, 15 and interaction with apoptotic cellderived vesicles 16 and exosomes. 17,18 Although the mechanisms have not been elucidated, it has been reported that DCs acquire proteins/peptides from other cells via exosomes. [17][18][19] Recently, it has been suggested that transfer of exosome-shuttle miRNAs might constitute a mechanism of cell-tocell communication that regulates mRNA translation 20 or, alternatively, a way to dispose of "unwanted" miRNAs. 21 An important unanswered question in the field is how exosome-shuttle miRNAs, transported inside the vesicles, are delivered into the cytosol of the acceptor cells, a problem we have investigated in this study with the use of DCs. Addressing this point has been challenging because (1) the composition of DC exosomes depends on the maturation of the DC of origin 22,23 ; (2) there is limited information on intercellular communication via "endogenous" (instead of exogenously added...

Recognition of injured mitochondria for degradation by macroautophagy is essential for cellular health, but the mechanisms remain poorly understood. Cardiolipin is an inner mitochondrial membrane phospholipid. We found that rotenone, staurosporine, 6-hydroxydopamine and other pro-mitophagy stimuli caused externalization of cardiolipin to the mitochondrial surface in primary cortical neurons and SH-SY5Y cells. RNAi knockdown of cardiolipin synthase or of phospholipid scramblase-3, which transports cardiolipin to the outer mitochondrial membrane, decreased mitochondrial delivery to autophagosomes. Furthermore, we found that the autophagy protein microtubule-associated-protein-1-light chain-3 (LC3), which mediates both autophagosome formation and cargo recognition, contains cardiolipin-binding sites important for the engulfment of mitochondria by the autophagic system. Mutation of LC3 residues predicted as cardiolipin-interaction sites by computational modeling inhibited its participation in mitophagy. These data indicate that redistribution of cardiolipin serves as an “eat-me” signal for the elimination of damaged mitochondria from neuronal cells.

Obesity increases the risk of adverse outcomes during acute critical illnesses such as burns, severe trauma, and acute pancreatitis. Although individuals with more body fat and higher serum cytokines and lipase are more likely to experience problems, the roles that these characteristics play are not clear. We used severe acute pancreatitis as a representative disease to investigate the effects of obesity on local organ function and systemic processes. In obese humans, we found that an increase in the volume of intrapancreatic adipocytes was associated with more extensive pancreatic necrosis during acute pancreatitis and that acute pancreatitis was associated with multisystem organ failure in obese individuals. In vitro studies of pancreatic acinar cells showed that unsaturated fatty acids were proinflammatory, releasing intracellular calcium, inhibiting mitochondrial complexes I and V, and causing necrosis. Saturated fatty acids had no such effects. Inhibition of lipolysis in obese (ob/ob) mice with induced pancreatitis prevented a rise in serum unsaturated fatty acids and prevented renal injury, lung injury, systemic inflammation, hypocalcemia, reduced pancreatic necrosis, and mortality. Thus, therapeutic approaches that target unsaturated fatty acid–mediated lipotoxicity may reduce adverse outcomes in obese patients with critical illnesses such as severe acute pancreatitis.

Reactivation of herpes simplex virus type 1 from neuronal latency is a common and potentially devastating cause of disease worldwide. CD8 + T cells can completely inhibit HSV reactivation in mice, with IFN-γ affording a portion of this protection. Here, we found that CD8 + T cell lytic granules are also required for the maintenance of neuronal latency both in vivo and in ex vivo ganglia cultures, and that their directed release to the junction with neurons in latently infected ganglia did not induce neuronal apoptosis. We describe a non-lethal mechanism of viral inactivation in which the lytic granule component, granzyme B, degrades the herpes simplex virus type 1 immediate early protein, ICP4, which is essential for further viral gene expression.Several lines of evidence support a role for CD8 + T cells in controlling herpes simplex virus type 1 (HSV-1) latency. CD8 + T cells, many expressing granzyme B (GrB), are found juxtaposed to HSV-1 latently infected sensory neurons of both humans (1-4) and mice (5-8). In C57BL/6 mice, CD8 + T cells specific for the immunodominant HSV-1 glycoprotein B 498-505 epitope (gB-CD8) polarize their T cell receptor (TCR) to junctions with neurons in situ forming apparent immunologic synapses (9). Murine gB-CD8 can block HSV-1 reactivation from latency in vivo and in ex vivo ganglia cultures in an MHC-dependent manner (9-11). Because HSV-1 establishes latency solely within ganglionic neurons (12,13), we hypothesize that some latently infected neurons directly present viral antigens to HSV-specific * This manuscript has been accepted for publication in Science. This version has not undergone final editing. Please refer to the complete version of record at http://www.sciencemag.org/. The manuscript may not be reproduced or used in any manner that does not fall within the fair use provisions of the Copyright Act without the prior, written permission of AAAS. Published version available at

GPD1-L is a novel gene that may affect trafficking of the cardiac Na+ channel to the cell surface. A GPD1-L mutation decreases SCN5A surface membrane expression, reduces inward Na+ current, and causes Brugada syndrome.