Regulatory T cells (T_regs) are found at sites of chronic inflammation where they mediate bystander and Ag-specific suppression of local immune responses. However, little is known about the molecular control of T_reg recruitment into inflamed human tissues. We report that up to 18% of T cells in areas of inflammation in human liver disease are forkhead family transcriptional regulator box P3 (FoxP3)⁺ T_regs. We isolated CD4⁺CD25⁺CD127^lowFoxP3⁺ T_regs from chronically inflamed human liver removed at transplantation; compared with blood-derived T_regs, liver-derived T_regs express high levels of the chemokine receptors CXCR3 and CCR4. In flow-based adhesion assays using human hepatic sinusoidal endothelium, T_regs used CXCR3 and α4β1 to bind and transmigrate, whereas CCR4 played no role. The CCR4 ligands CCL17 and CCL22 were absent from healthy liver, but they were detected in chronically inflamed liver where their expression was restricted to dendritic cells (DCs) within inflammatory infiltrates. These DCs were closely associated with CD8 T cells and CCR4⁺ T_regs in the parenchyma and septal areas. Ex vivo, liver-derived T_regs migrated to CCR4 ligands secreted by intrahepatic DCs. We propose that CXCR3 mediates the recruitment of T_regs via hepatic sinusoidal endothelium and that CCR4 ligands secreted by DCs recruit T_regs to sites of inflammation in patients with chronic hepatitis. Thus, different chemokine receptors play distinct roles in the recruitment and positioning of T_regs at sites of hepatitis in chronic liver disease.

The common lymphatic endothelial and vascular endothelial receptor (CLEVER-1; also known as FEEL-1 and stabilin-1) is a recycling and intracellular trafficking receptor with multifunctional properties. In this study, we demonstrate increased endothelial expression of CLEVER-1/stabilin-1 at sites of leukocyte recruitment to the inflamed human liver including sinusoids, septal vessels, and lymphoid follicles in inflammatory liver disease and tumor-associated vessels in hepatocellular carcinoma. We used primary cultures of human hepatic sinusoidal endothelial cells (HSEC) to demonstrate that CLEVER-1/stabilin-1 expression is enhanced by hepatocyte growth factor but not by classical proinflammatory cytokines. We then showed that CLEVER-1/stabilin-1 supports T cell transendothelial migration across HSEC under conditions of flow with strong preferential activity for CD4 FoxP3⁺ regulatory T cells (Tregs). CLEVER-1/stabilin-1 inhibition reduced Treg transendothelial migration by 40% and when combined with blockade of ICAM-1 and vascular adhesion protein-1 (VAP-1) reduced it by >80%. Confocal microscopy demonstrated that 60% of transmigrating Tregs underwent transcellular migration through HSEC via ICAM-1– and VAP-1–rich transcellular pores in close association with CLEVER-1/stabilin-1. Thus, CLEVER-1/stabilin-1 and VAP-1 may provide an organ-specific signal for Treg recruitment to the inflamed liver and to hepatocellular carcinoma.

Chemokines direct leukocyte trafficking and positioning within tissues, thus playing critical roles in regulating immune responses and inflammation. The chemokine system is complex, involving interactions between multiple chemokines and their receptors that operate in combinatorial cascades with adhesion molecules. The involvement of multiple chemokines and chemokine receptors in these processes brings flexibility and specificity to recruitment. The hepatic vascular bed is a unique low-flow environment through which leukocytes are recruited to the liver during homeostatic immune surveillance and in response to infection or injury. The rate of leukocyte recruitment and the nature of cells recruited through the sinusoids in response to inflammatory signals will shape the severity of disease. At one end of the spectrum, fulminant liver failure results from a rapid recruitment of leukocytes that leads to hepatocyte destruction and liver failure; at the other end, diseases such as chronic hepatitis C infection may progress over many years from hepatitis to fibrosis and cirrhosis. Chronic hepatitis is characterized by a T lymphocyte-rich infiltrate and the nature and outcome of hepatitis will depend on the T cell subsets recruited, their activation and function within the liver. Different subsets of effector T cells have been described based on their secretion of cytokines and specific functions. These include Th1 and Th2 cells, and more recently Th17 and Th9 cells, which are associated with different types of immune response and which express distinct patterns of chemokine receptors that promote their recruitment under particular conditions. The effector function of these cells is balanced by the recruitment of regulatory T cells that are able to suppress antigen-specific effectors to allow resolution of immune responses and restoration of immune homeostasis. Understanding the signals that are responsible for recruiting different lymphocyte subsets to the liver will elucidate disease pathogenesis and open up new therapeutic approaches to modulate recruitment in favor of resolution rather than injury.

Carcinogenesis on the background of liver fibrosis: Implications for the management of hepatocellular cancer

Liver sinusoidal endothelial cells (LSEC) form a unique barrier between the liver sinusoids and the underlying parenchyma, and thus play a crucial role in maintaining metabolic and immune homeostasis, as well as actively contributing to disease pathophysiology. Whilst their endocytic and scavenging function is integral for nutrient exchange and clearance of waste products, their capillarisation and dysfunction precedes fibrogenesis. Furthermore, their ability to promote immune tolerance and recruit distinct immunosuppressive leukocyte subsets can allow persistence of chronic viral infections and facilitate tumour development. In this review, we present the immunological and barrier functions of LSEC, along with their role in orchestrating fibrotic processes which precede tumourigenesis. We also summarise the role of LSEC in modulating the tumour microenvironment, and promoting development of a pre-metastatic niche, which can drive formation of secondary liver tumours. Finally, we summarise closely inter-linked disease pathways which collectively perpetuate pathogenesis, highlighting LSEC as novel targets for therapeutic intervention.

Significance: Organ fibrosis is a major cause of global morbidity and mortality. It is driven by chronic inflammation and associated oxidative stress with depletion of cellular antioxidant defenses. We demonstrate a pathway in which the evolutionarily conserved receptor stabilin-1 on tissue-infiltrating macrophages provides a second-line defense to prevent tissue damage from oxidative stress. Stabilin-1 ⁺ monocytes take up malondialdehyde-LDL (MDA-LDL), a major product of oxidative lipid peroxidation, to form ceroid-laden macrophages. Through the uptake of MDA-LDL, stabilin-1 suppresses production of the profibrogenic chemokine CCL3 and prevents excessive collagen deposition in experimental models of liver fibrosis. We propose that macrophage stabilin-1 is a critical defense against oxidative tissue damage and thereby maintains tissue homeostasis.