Macrophages and derived-TNF-α promote lipopolysaccharide-induced upregulation of endogenous β-glucuronidase in the epithelial cells of the bile duct: A possible facilitator of hepatolithiasis formation

Hepatolithiasis is prevalent in Southeast Asian regions, and the role of endogenous β-glucuronidase (β-GD) in the formation of hepatolithiasis is being gradually recognised. Revealing the regulation mechanism of the expression of endogenous β-GD will provide new therapeutic strategies for intervening in the formation of hepatolithiasis.

Infiltration of macrophages, especially M2 macrophages, may be involved in the hepatolithiasis formation. LPS activates the macrophages, inducing the secretion of TNF-α, which can further increase the expression of endogenous β-GD in the epithelial cells of the bile duct, possibly via the NF-κB/PCNA signalling cascade.

Liver specimens from patients with hepatolithiasis were examined by immunohistochemistry to assess the expression of macrophage markers including CD68, CD80, and CD206, as well as that of TNF-α and endogenous β-GD, compared with that in normal liver samples. HiBEpiC cells were co-cultured directly or indirectly with induced M2 macrophages or directly stimulated with TNF-α, and the expression of the endogenous β-GD was examined. A PKC inhibitor, chelerythrine, and an NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), were used to elucidate the possible regulation mechanism.

The expression of macrophage markers including CD68 and CD206, as well as that of TNF-α and endogenous β-GD significantly increased in liver specimens from patients with hepatolithiasis compared with that in normal liver samples. The expression of CD68, CD206 and TNF-α was positively correlated with that of endogenous β-GD. When HiBEpiC cells were co-cultured directly or indirectly with M2 macrophages, following stimulation with lipopolysaccharide (LPS), the expression of endogenous β-GD was significantly higher in the indirect co-culture group than that in the direct co-culture group, or in HiBEpiC cells or M2 macrophages cultured alone. Further experiments revealed that following stimulation with LPS, TNF-α secretion increased in both the indirect and direct co-culture groups compared with that in HiBEpiC cells cultured alone. TNF-α increased the expression of endogenous β-GD in HiBEpiC cells, in a dose- and time-dependent manner. In addition, TNF-α significantly increased the expression levels of p-P65 and proliferating cell nuclear antigen (PCNA), and PDTC effectively inhibited the TNF-α-induced expression of PCNA and β-GD. Conclusions: Infiltration of macrophages, especially M2 macrophages, may be involved in the hepatolithiasis formation. LPS activates the macrophages, inducing the secretion of TNF-α, which can further increase the expression of endogenous β-GD in the epithelial cells of the bile duct, possibly via the NF-κB/PCNA signalling cascade.