Irina Gurevich

Luminal Bacteria Recruit CD103+ Dendritic Cells into the Intestinal Epithelium to Sample Bacterial Antigens for Presentation

CD103+ dendritic cells (DCs) carry bacteria from the small intestine and can present antigens to T cells. Yet they have not been recorded sampling luminal bacteria or presenting bacterial antigens in mesentery lymph nodes. We used 2-photon microscopy in live Cx3cr1(+/gfp) ×Cd11c-YFP mice to study these processes. At steady state, sparse CD103+ DCs occupied the epithelium. They patrolled among enterocytes while extending dendrites toward the lumen, likely using tight-junction proteins to penetrate the epithelium. Challenge with Salmonella triggered chemokine- and toll-like receptor (TLR)-dependent recruitment of additional DCs from the lamina propria (LP). The DCs efficiently phagocytosed the bacteria using intraepithelial dendrites. Noninvasive bacteria were similarly sampled. In contrast, CD103+ DCs sampled soluble luminal antigen inefficiently. In mice harboring CD103+ DCs, antigen-specific CD8 T cells were subsequently activated in MLNs. Intestinal CD103+ DCs are therefore equipped with unique mechanisms to independently complete the processes of uptake, transportation, and presentation of bacterial antigens.

DC mobilization from the skin requires docking to immobilized CCL21 on lymphatic endothelium and intralymphatic crawling

Dermal DC mobilization requires docking to CCL21 on lymphatic endothelium

Models of Motor Adaptation and Impedance Control in Human Arm Movements

Abstract Earlier studies of human reaching movements have suggested that the arm trajectory formation processes are hierarchically organized: higher levels deal with the planning of desired motions for the hand while lower levels take care of their execution by taking advantage of the viscoelastic properties of muscles. Here we review recent studies of motor adaptation to elastic loads aimed at examining the validity of this two-stage control scheme. Movement and postural stiffness field measurements were conducted both in the unloaded and loaded cases. The observed behavior indicated that following only a few practice trials in the presence of the load, the loaded movements tended to converge toward straight hand paths with bell-shaped velocity profiles adequately described by the minimum-jerk model. A model was developed which suggests that load adaptation involves the tuning of both the arm impedance parameters and the hand equilibrium trajectory and that the new equilibrium trajectory and stiffness field result from the summation of posture-related components needed to overcome the load and “phasic” components responsible for driving the arm along the desired trajectory. Computer simulations demonstrated that the proposed adaptation scheme can successfully account for the motor behavior observed both before and after the application of the loads. The implications of the proposed scheme are discussed in view of recent neurophysiological studies of unloaded and loaded reaching movements.

The integrin coactivator Kindlin-3 is not required for lymphocyte diapedesis

Kindlin-3 is an integrin-binding focal adhesion adaptor absent in patients with leukocyte and platelet adhesion deficiency syndrome and is critical for firm integrin-dependent leukocyte adhesion. The role of this adaptor in leukocyte diapedesis has never been investigated. In the present study, the functions of Kindlin-3 in this process were investigated in effector T lymphocytes trafficking to various lymphoid and nonlymphoid tissues. In vitro, Kindlin-3-deficient T cells displayed severely impaired lymphocyte function antigen-1-dependent lymphocyte adhesion but partially conserved very late antigen-4 adhesiveness. In vivo, the number of adoptively transferred Kindlin-3-deficient T effectors was dramatically elevated in the circulating pool compared with normal effectors, and the Kindlin-3 mutant effectors failed to enter inflamed skin lesions. The frequency of Kindlin-3-deficient T effectors arrested on vessel walls within inflamed skin-draining lymph nodes was also reduced. Strikingly, however, Kindlin-3-deficient effector T cells accumulated inside these vessels at significantly higher numbers than their wild-type lymphocyte counterparts and successfully extravasated into inflamed lymph nodes. Nevertheless, on entering these organs, the interstitial motility of these lymphocytes was impaired. This is the first in vivo demonstration that Kindlin-3-stabilized integrin adhesions, although essential for lymphocyte arrest on blood vessels and interstitial motility, are not obligatory for leukocyte diapedesis.

Hyaluronan synthesis is necessary for autoreactive T-cell trafficking, activation, and Th1 polarization

Significance 4-methylumbelliferone (4-MU) is an oral drug that inhibits synthesis of hyaluronan, an extracellular matrix polymer implicated in autoimmunity. In the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS), 4-MU drives the polarization of T cells away from a Th1 phenotype, associated with disease progression, and toward a FoxP3+ regulatory T-cell phenotype, associated with disease prevention. Moreover, 4-MU inhibits the reactive response of astrocytes, immunocompetent resident cells of the CNS, and prevents trafficking of activated T cells to the CNS. These data suggest that the extracellular matrix, and hyaluronan in particular, may be an active contributor to autoimmune pathogenesis in EAE and, furthermore, that 4-MU is a promising strategy for treatment of CNS autoimmunity. The extracellular matrix polysaccharide hyaluronan (HA) accumulates at sites of autoimmune inflammation, including white matter lesions in multiple sclerosis (MS), but its functional importance in pathogenesis is unclear. We have evaluated the impact of 4-methylumbelliferone (4-MU), an oral inhibitor of HA synthesis, on disease progression in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS. Treatment with 4-MU decreases the incidence of EAE, delays its onset, and reduces the severity of established disease. 4-MU inhibits the activation of autoreactive T cells and prevents their polarization toward a Th1 phenotype. Instead, 4-MU promotes polarization toward a Th2 phenotpye and induction of Foxp3+ regulatory T cells. Further, 4-MU hastens trafficking of T cells through secondary lymphoid organs, impairs the infiltration of T cells into the CNS parenchyma, and limits astrogliosis. Together, these data suggest that HA synthesis is necessary for disease progression in EAE and that treatment with 4-MU may be a potential therapeutic strategy in CNS autoimmunity. Considering that 4-MU is already a therapeutic, called hymecromone, that is approved to treat biliary spasm in humans, we propose that it could be repurposed to treat MS.

Arm Stiffness And Movement Adaptation To External Loads

In this study we have combined kinematic recording and analysis of movement trajectories with computer simulations, based on the equilibrium trajectory control scheme, in order to investigate what mechanisms underlie motor adaptation to unexpectedly introduced external loads. The static arm stiffness field while maintaining posture against externally imposed forces was measured and a mathematical model to account for the variations in joint stiffnesses with external forces was developed. Our results have indicated that the observed motor behavior in the loaded case can be well accounted for within the framework of the equilibrium trajectory control hypothesis.

Murine anti–third-party central-memory CD8+ T cells promote hematopoietic chimerism under mild conditioning: lymph-node sequestration and deletion of anti-donor T cells

Transplantation of T cell-depleted BM (TDBM) under mild conditioning, associated with minimal toxicity and reduced risk of GVHD, offers an attractive therapeutic option for patients with nonmalignant hematologic disorders and can mediate immune tolerance to subsequent organ transplantation. However, overcoming TDBM rejection after reduced conditioning remains a challenge. Here, we address this barrier using donorderived central memory CD8(+) T cells (Tcms), directed against third-party antigens. Our results show that fully allogeneic or (hostXdonor)F1-Tcm, support donor chimerism (> 6 months) in sublethally irradiated (5.5Gy) mice, without GVHD symptoms. Chimerism under yet lower irradiation (4.5Gy) was achieved by combining Tcm with short-term administration of low-dose Rapamycin. Importantly, this chimerism resulted in successful donor skin acceptance, whereas third-party skin was rejected. Tracking of host anti-donor T cells (HADTCs), that mediate TDBMT rejection, in a novel bioluminescence-imaging model revealed that Tcms both induce accumulation and eradicate HADTCs in the LNs,concomitant with their elimination from other organs, including the BM. Further analysis with 2-photon microcopy revealed that Tcms form conjugates with HADTCs, resulting in decelerated and confined movement of HADTCs within the LNs in an antigen-specific manner. Thus, anti-third-party Tcms support TDBMT engraftment under reduced-conditioning through lymph-node sequestration and deletion of HADTCs, offering a novel and potentially safe approach for attaining stable hematopoietic chimerism.

Heparanase of murine effector lymphocytes and neutrophils is not required for their diapedesis into sites of inflammation

Heparanase, the exclusive mammalian heparan sulfate‐degrading enzyme, has been suggested to be utilized by leukocytes to penetrate through the dense basement membranes surrounding blood venules. Despite its established role in tumor cell invasion, heparanase function in leukocyte extravasation has never been demonstrated. We found that TH1/TC1‐type effector T cells are highly enriched for this enzyme, with a 3.6‐fold higher heparanase mRNA expression compared with naive lymphocytes. Using adoptive transfer of wild‐type and heparanase‐deficient effector T cells into inflamed mice, we show that T‐cell heparanase was not required for extravasation inside inflamed lymph nodes or skin. Leukocyte extravasation through acute inflamed skin vessels was also heparanase independent. Furthermore, neutrophils emigrated to the inflamed peritoneal cavity independently of heparanase expression on either the leukocytes or on the endothelial and mesothelial barriers, and overexpression of the enzyme on neutrophils did not facilitate their emigration. However, heparanase absence significantly reduced monocyte emigration into the inflamed peritoneal cavity. These results collectively suggest that neither leukocyte nor endothelial heparanase is required for T‐cell and neutrophil extravasation through inflamed vascular barriers, whereas this enzyme is required for optimal monocyte recruitment to inflamed peritoneum.—Stoler‐Barak, L., Petrovich, E., Aychek, T., Gurevich, I., Tal, O., Hatzav, M., Ilan, N., Feigelson, S. W., Shakhar, G., Vlodavsky, I., Alon, R. Heparanase of murine effector lymphocytes and neutrophils is not required for their diapedesis into sites of inflammation. FASEB J. 29, 2010‐2021 (2015). www.fasebj.org

Active dissemination of cellular antigens by DCs facilitates CD8+ T‐cell priming in lymph nodes

Antigen (Ag) specific activation of naïve T cells by migrating dendritic cells (DCs) is a highly efficient process, although the chances for their colocalization in lymph nodes (LNs) appear low. Ag presentation may be delegated from Ag‐donor DCs to the abundant resident DCs, but the routes of Ag transfer and how it facilitates T‐cell activation remain unclear. We visualized CD8+ T cell‐DC interactions to study the sites, routes, and cells mediating Ag transfer in mice. In vitro, Ag transfer from isolated ovalbumin (OVA)+ bone marrow (BM) DCs triggered widespread arrest, Ca2+ flux, and CD69 upregulation in OT‐I T cells contacting recipient DCs. Intravital two‐photon imaging revealed that survival of Ag‐donor DCs in LNs was required for Ag dissemination among resident CD11c+ DCs. Upon interaction with recipient DCs, CD8+ T cells clustered, upregulated CD69, proliferated and differentiated into effectors. Few DCs sufficed for activation, and for efficient Ag dissemination lymphocyte function associated antigen 1 (LFA‐1) expression on recipient DCs was essential. Similar findings characterized DCs infected with a replication‐deficient OVA‐expressing Vaccinia virus known to downregulate MHC‐I. Overall, active Ag dissemination from live incoming DCs helped activate CD8+ T cells by increasing the number of effective presenting cells and salvaged T‐cell priming when Ag‐donor DCs could not present Ag.

The bone marrow is patrolled by NK cells that are primed and expand in response to systemic viral activation.

The bone marrow hosts NK cells whose distribution, motility and response to systemic immune challenge are poorly understood. At steady state, two‐photon microscopy of the bone marrow in Ncr1gfp/+ mice captured motile NK cells interacting with dendritic cells. NK cells expressed markers and effector molecules of mature cells. Following poly (I:C) injection, RNA‐Seq of NK cells revealed three phases of transcription featuring immune response genes followed by posttranscriptional processes and proliferation. Functionally, poly (I:C) promoted upregulation of granzyme B, enhanced cytotoxicity in vitro and in vivo, and, in the same individual cells, triggered proliferation. Two‐photon imaging revealed that the proportion of sinusoidal NK cells decreased, while at the same time parenchymal NK cells accelerated, swelled and divided within the bone marrow. MVA viremia induced similar responses. Our findings demonstrate that the bone marrow is patrolled by mature NK cells that rapidly proliferate in response to systemic viral challenge while maintaining their effector functions.