The G protein coupled bile acid receptor TGR5 (Gpbar1) modulates endothelin-1 signaling in liver

Introduction:

TGR5 (Gpbar-1) is a G-Protein-coupled bile acid (BA) receptor (Kawamata et al. 2003) expressed on nonparenchymal liver cells such as hepatic stellate cells (HSCs) and liver sinusoidal endothelial cells (LSECs) (Keitel et al. 2006, 2008). Lithocholic acid (LCA) is a hydrophobic bile acid. Feeding mice with a diet containing 1%LCA leads to the development of a severe toxic liver injury (Fickert et al. 2006). The most relevant complication of liver damage is the development of portal hypertension (PH). It has been demonstrated previously that activated HSCs contribute to PH by Endothelin-1 (ET-1)-induced cell-contraction in a cAMP-dependent manner (Rockey et al 1993, Reinehr et al. 2002). If TGR5 plays a role during this process was unknown.

Methods:

8 – 12 week old wildtype (WT) and TGR5 knockout (KO) mice were sacrificed after receiving a diet containing 1%LCA for 84 hrs. Liver damage was determined via analysis of liver enzymes in serum and HE-staining. Portal pressure was measured by cannulation of the portal vein. Hepatic mRNA-expression of different genes was determined by RT-PCR analysis. HSC contraction was tested using an HSC-contraction assay. Localization of the ETA-receptor on HSCs was analyzed by immunofluorescence (IF) and colocalization-analysis. ET-1 levels in LSECs were measured via ELISA-immunoassay and RT-PCR.

Results:

TGR5 KO mice suffered from a more severe liver damage as compared to WT animals when fed a 1%LCA diet as determined by the area of necrosis seen in liver tissue as well as increased serum AST-levels. Measurement of the portal pressure in these mice revealed an induction of PH in LCA-fed KO mice but in none of the control groups. RT-PCR-analysis confirmed an increased expression of genes associated with PH, e.g. ET-1 in livers of LCA-fed TGR5 KO mice. Challenge of LSECs as the most important source of ET-1 in the liver with a TGR5 agonist caused a significant reduction of ET-1 secretion. Moreover, treatment of mice and isolated HSCs with a TGR5 agonist was able to attenuate ET-1-induced PH as well as HSC contraction, respectively. By means of IF of the ETA-receptor on the surface of HSCs and colocalization analysis we were able to demonstrate internalization of this receptor after treatment with a TGR5 agonist.

Conclusion:

In conclusion, TGR5 plays a protective role against the development of PH after induction of liver injury by two synergistically acting mechanisms.