Fractalkine and other chemokines in primary biliary cirrhosis

Abstract

Primary biliary cirrhosis (PBC) is characterized by the autoimmune injury of small intrahepatic bile duct. On this basis, it has been suggested that the targeted biliary epithelial cells (BEC) play an active role in the perpetuation of autoimmunity by attracting immune cells via chemokine secretion. To address this issue, we challenged BEC using multiple toll-like receptor (TLR) ligands as well as autologous liver infiltrating mononuclear cells (LMNC) with subsequent measurement of BEC phenotype and chemokine production and LMNC chemotaxis by quantifying specific chemokines, specially CX3CL1 (fractalkine). We submit the hypothesis that BEC are in fact the innocent victims of the autoimmune injury and that the adaptive immune response is critical in PBC.

Figure 1

Chemokines produced by biliary epithelial cells under basal conditions or after stimulation with TLR3 ligand (poly I : C) for 48 hours. Cell-free culture supernatants were analyzed by a protein array kit to evaluate 174 different proteins simultaneously. Unstimulated cells produced detectable amounts of GRO-α/CXCL1, ENA-78/CXCL5, GCP-2/CXCL6, and IL-8/CXCL8, while poly I : C stimulation led to enhanced MIP-1α/CCL3, MIP-1β/CCL4, RANTES/CCL5, and IP-10/CXCL10.

Figure 2

(a) Fractalkine production from endothelial cells from PBC and control (chronic hepatitis B and C) livers exposed to TLR ligands. Endothelial cells produced fractalkine with LTA, poly I : C, LPS, and flagellin with no significant differences observed between patients and control livers. (b) BEC did not produce fractalkine with any additional TLR ligand.

Figure 3

Autologous liver mononuclear cells adhesion assay using endothelial cells and BEC after stimulation with TLR4 ligand (LPS). Adherent liver mononuclear cells were stained and counted in ten random high-power microscopy fields. Liver mononuclear cells from PBC livers adhered in greater numbers than did liver mononuclear cells from controls using either endothelial cells or biliary epithelial cells, whereas liver mononuclear cells adhered only minimally to liver sinusoidal endothelial cells in all instances. Other TLR ligands did not accelerate liver mononuclear cells adhesion with neither endothelial cells nor biliary epithelial cells (data not shown).

Figure 4

The proposed role of fractalkine is illustrated. TLR3 or TLR4 ligands stimulate vascular endothelial cells to produce fractalkine as chemokine, then fractalkine attracts CX3CR1 positive monocytes or NK cells. Subsequently, TLR4 ligand stimulated BEC produce fractalkine as cell adhesion molecule, then fractalkine recruit CX3CR1 positive cells around PBC target cells. This starts the chronic nonsuppurative destructive cholangitis and perpetuates the autoimmune pathogenesis of disease. Finally, TLR3 ligand stimulated biliary epithelial cells produce Th1 chemokines, and these chemokines are considered to contribute this autoimmune mechanism.