Elucidating cellular mechanisms that maintain the intrahepatic immune balance is crucial to our understanding of viral or autoimmune liver diseases and allograft acceptance. Liver sinusoidal endothelial cells (LSECs) play an important role in modifying local immune responses to tolerance in major histocompatibility complex (MHC) I-restricted models, whereas their contribution in the MHCII context is still controversial. In an MHCII chimeric mouse model that excludes MHCII-mediated antigen presentation by professional antigen-presenting cells, we demonstrated that LSECs prime CD4 ؉ T cells to a CD45RB low memory phenotype lacking marker cytokine production for effector cells that was stable in vivo following immunogenic antigen re-encounter. Although these cells, which we term T LSEC , had the capacity to enter lymph nodes and the liver, they did not function as effector cells either in a delayed-type hypersensitivity reaction or in a hepatitis model. T LSEC inhibited the proliferation of naïve CD4 ؉ T cells in vitro although being CD25 low and lacking expression of forkhead box protein (FoxP)3. Furthermore, these cells suppressed hepatic inflammation as monitored by alanine aminotransferase levels and cellular infiltrates in a T cell-mediated autoimmune hepatitis model in vivo. Conclusion: T LSEC first described here might belong to the expanding group of FoxP3 ؊ regulatory T cells. Our findings strengthen the previously discussed assumption that CD4 ؉ T cell priming by nonprofessional antigen-presenting cells induces anti-inflammatory rather than proinflammatory phenotypes. Because recruitment of CD4 ؉ T cells is increased upon hepatic inflammation, T LSEC might contribute to shifting antigen-dependent immune responses to tolerance toward exogenous antigens or toward endogenous self-antigens, especially under inflammatory conditions. (HEPATOLOGY 2009;50:1904-1913 U nder physiological conditions, the liver seems rather to support the induction of peripheral tolerance than to establish immunity. 1 In particular, liver sinusoidal endothelial cells (LSECs) are believed to shift the hepatic immune balance toward tolerance by presenting major histocompatibility complex (MHC) Irestricted antigens, thereby inducing anergy and tolerance. 2,3 LSECs constitutively express MHCII and costimulatory molecules 4 and take up exogenous antigens. 2 Naïve CD4 ϩ T cells migrate into the liver 5,6 and interact with LSECs. 7 This raises the question whether LSECs also prime CD4 ϩ T cells. Coculturing naïve CD4 ϩ T cells with LSECs resulted in stimulation of the T cells. 8 Another recent study using LSEC preparations depleted of cells of hematopoietic origin has created doubts about the priming capacity of LSECs for naïve CD4 ϩ T cells in the absence of professional antigenpresenting cells (APCs). 9 Priming of naïve CD4 ϩ T cells by other nonprofessional APCs such as aortic endothelial cells or naïve B cells induces distinct T cell populations lacking cytokine production and displaying suppressive capaci-

The pathogenesis of autoimmune liver diseases is poorly understood. Animal models are necessary to investigate antigen presentation and priming of T-cells in the context of autoimmunity in the liver. Transgenic mouse models were generated in which the model antigen ovalbumin is expressed in hepatocytes (TF-OVA) or cholangiocytes (ASBT-OVA). A utoimmune hepatitis and cholangitis are triggered by autoreactive T-cells. Animal models are needed to study the early events in their pathogenesis, namely, the priming of autoreactive T-cells. The requirements for an immune-based animal model are restriction of the immune response to the liver, antigen specificity, and the potential to study the priming of CD8 and CD4 T-cells. Several animal models of autoimmune hepatitis have been developed, 1 none of which fulfills these criteria. Transgenic expression of foreign major histocompatibility complex (MHC) class I molecules in the liver has been widely used. [2][3][4] However, this model has a significant disadvantage: unlike the pathophysiology of an immune or autoimmune reaction, CD8 T cells recognize their antigen on hepatocytes but the antigen is not presented by professional antigen-presenting cells (APCs) in this model. Injection of the synthetic peptide ova p257-264 (SIINFEKL) into mice followed by transfer of antigenspecific CD8 T-cells has also been used, 5 but the possibility of peptide binding to MHC-I molecules on T-cells themselves and their activation by each other 6 or by unrelated cells is a possible concern. Infection with virus expressing ovalbumin 7 results in temporary expression of ovalbumin but also activation of innate immune mechanisms. No animal model exists that allows the simulta-

No abstract

Gut-activated T cells migrating into the liver can cause extraintestinal manifestations of inflammatory bowel disease. T cells acquire a gut-homing phenotype dependent on retinoic acid (RA) provided by intestinal dendritic cells (DC). We investigated whether liver antigen-presenting cells can induce gut tropism supporting an enterohepatic lymphocyte circulation. Priming of CD4+ T cells by liver sinusoidal endothelial cells (LSEC) supported migration into gut and gut-associated lymphoid tissue. As observed for T cells primed by intestinal DCs, this gut tropism depended on α4β7 integrin and CC chemokine receptor 9 (CCR9) expression by LSEC-primed CD4+ T cells. The induction of gut-homing molecules was mediated by RA, a derivate of vitamin A that is stored in large amounts within the liver. LSECs expressed functional retinal dehydrogenases and could convert vitamin A to RA. Conversely, the lack of signaling via the RA receptor prevented the expression of α4β7 integrin and CCR9 on LSEC-primed CD4+ T cells, consequently reducing their in vivo migration to the intestine. Other liver antigen-presenting cells failed to support high expression of α4β7 integrin on CD4+ T cells, thus, the potential to induce gut homing is restricted to LSECs. Conclusion: The capacity to promote gut tropism via vitamin A use is not unique for intestinal DCs but is also a feature of LSECs. Our data support the assumption that CD4+ T cells can migrate from the liver to the gut as one branch of a postulated enterohepatic lymphocyte circulation.

Leukocyte adhesion and transmigration are central features governing immune surveillance and inflammatory reactions in body tissues. Within the liver sinusoids, chemokines initiate the first crucial step of T-cell migration into the hepatic tissue. We studied molecular mechanisms involved in endothelial chemokine supply during hepatic immune surveillance and liver inflammation and their impact on the recruitment of CD4+ T cells into the liver. In the murine model of Concanavalin A-induced T cell-mediated hepatitis, we showed that hepatic expression of the inflammatory CXC chemokine ligands (CXCL)9 and CXCL10 strongly increased whereas homeostatic CXCL12 significantly decreased. Consistently, CD4+ T cells expressing the CXC chemokine receptor (CXCR)3 accumulated within the inflamed liver tissue. In histology, CXCL9 was associated with liver sinusoidal endothelial cells (LSEC) which represent the first contact site for T-cell immigration into the liver. LSEC actively transferred basolaterally internalized CXCL12, CXCL9 and CXCL10 via clathrin-coated vesicles to CD4+ T cells leading to enhanced transmigration of CXCR4+ total CD4+ T cells and CXCR3+ effector/memory CD4+ T cells, respectively in vitro. LSEC-expressed CXCR4 mediated CXCL12 transport and blockage of endothelial CXCR4 inhibited CXCL12-dependent CD4+ T-cell transmigration. In contrast, CXCR3 was not involved in the endothelial transport of its ligands CXCL9 and CXCL10. The clathrin-specific inhibitor chlorpromazine blocked endothelial chemokine internalization and CD4+ T-cell transmigration in vitro as well as migration of CD4+ T cells into the inflamed liver in vivo. Moreover, hepatic accumulation of CXCR3+ CD4+ T cells during T cell-mediated hepatitis was strongly reduced after administration of chlorpromazine. These data demonstrate that LSEC actively provide perivascularly expressed homeostatic and inflammatory chemokines by CXCR4- and clathrin-dependent intracellular transport mechanisms thereby contributing to the hepatic recruitment of CD4+ T-cell populations during immune surveillance and liver inflammation.