The role of PGE2 biosynthesis and metabolism in liver injury and liver cancer
Description
PGE2 plays an important role in liver inflammation and carcinogenesis. Its metabolism is regulated by a cascade of reactions catalyzed by enzymes including COX-1/2, mPGES-1/2, 15-PGDH. Among these regulators, mPGES-1 is a cytokine-inducible enzyme mainly responsible for catalyzing terminal synthesis of PGE2, 15-PGDH catalyzes the oxidation of PGE2 to 15-keto-PGE2. In this context, we exogenously expressed mPGES-1 or 15-PGDH genes in mice hepatocytes to constitute a physiological condition ideal for evaluating PGE2 and its metabolites function in liver pathogenesis. In the first part, we developed transgenic mice with targeted expression of mPGES-1 in the liver and assessed the response of the transgenic mice to Fas-induced hepatocyte apoptosis and acute liver injury. Compared to wild type mice, the mPGES-1 Tg mice showed less liver hemorrhage, lower serum alanine transaminase and aspartate transaminase levels, less hepatic necrosis/apoptosis, and lower levels of caspase activation after intraperitoneal injection of the anti-Fas antibody Jo2. Western blotting analyses revealed increased expression and activation of the serine/threonine kinase Akt and associated anti-apoptotic molecules in the liver tissues of Jo2-treated mPGES-1 Tg mice. Pretreatment with the mPGES-1 inhibitor (MF63) or the Akt inhibitor (Akt inhibitor V) restored the susceptibility of the mPGES-1 Tg mice to Fas-induced liver injury. Our findings provide novel evidence that mPGES-1 prevents Fas-induced liver injury through activation of Akt and related signaling. This finding is consistent with previous reports of the anti-apoptotic and pro-proliferative role of PGE2. Our results suggest that induction of mPGES-1 or treatment with PGE2 may represent a potential therapeutic strategy for the prevention and treatment of Fas-associated liver injuries. In the second part, we generated transgenic mice with targeted expression of 15-PGDH in the liver and the animals were subjected to LPS/GalN-induced acute liver inflammation and injury. Compared to the wild type mice, the 15-PGDH Tg mice showed lower levels of alanine aminotransferase and aspartate aminotransferase, less liver tissue damage, less hepatic apoptosis/necrosis, less macrophage activation, and lower inflammatory cytokine production. In Kupffer cell cultures, treatment with 15-keto-PGE2 or the conditioned medium (CM) from 15-PGDH Tg hepatocyes inhibited LPS-induced cytokine production. Both 15-keto-PGE2 and the CM from15-PGDH Tg hepatocyes also up-regulated the expression of PPAR-γ downstream genes in Kupffer cells. In cultured hepatocytes, 15-keto-PGE2 treatment or 15-PGDH overexpression did not influence TNF-α-induced hepatocyte apoptosis. These findings suggest that 15-PGDH protects against LPS/GalN-induced liver injury and the effect is mediated via 15-keto-PGE2, which activates PPAR-γ in Kupffer cells and thus inhibits their ability to produce inflammatory cytokines. Accordingly, we observed that the PPAR-γ antagonist, GW9662, reversed the effect of 15-keto-PGE2 in Kupffer cell in vitro and restored the susceptibility of 15-PGDH Tg mice to LPS/GalN-induced acute liver injury in vivo. Our findings not only support the pro-inflammatory role of PGE2, but also reveal a novel anti-inflammatory role of 15-keto-PGE2. The data suggest that induction of 15-PGDH expression or utilization of a 15-keto-PGE2 analog may be therapeutic for treatment of endotoxin-associated liver inflammation/injury. Consistent with a pro-carcinogenic role for PGE2, overexpression mPGES-1 enhances growth of either HCC or cholangiocarcinoma cells, while overexpression 15-PGDH inhibits tumor cell growth in vitro. In the third part, we use a pharmacological method to induce 15-PGDH in cholangiocarcinoma tumor cells to inhibit PGE2 production. Our results indicated that treatment of human cholangiocarcinoma cells (CCLP1 and TFK-1) with ω-3 PUFA (DHA) or transfection of these cells with the Fat-1 gene (encoding Caenorhabditis elegans desaturase which converts ω-6 PUFA to ω-3 PUFA) significantly increased 15-PGDH protein level in cholangiocarcinoma cell lines. Human cholangiocarcinoma cells treated with DHA or transfected with a Fat-1 expression vector showed reduction of miRNA26a and miRNA26b (both miRNAs target 15-PGDH mRNA thus inhibiting 15-PGDH translation). Consistent with these findings, we observed that overexpression of miR26a or miR26b decreased 15-PGDH protein, reversed ω-3 PUFA-induced accumulation of 15-PGDH protein, and prevented ω-3 PUFA-induced inhibition of cholangiocarcinoma cell growth. Knockdown of 15-PGDH also attenuated ω-3 PUFA-induced inhibition of tumor cell growth. We observed that ω-3 PUFA suppressed miRNA26a and miRNA26b by inhibiting c-myc, a transcription factor that co-regulates a gene cluster comprised of miR-26a/b and carboxy-terminal domain RNA polymerase II polypeptide A small phosphatases (CTDSPs). Accordingly, overexpression of c-myc enhanced the expression of miRNA26a/b and prevented ω-3 PUFA-induced inhibition of tumor cell growth. Taken together, our results support a pro-tumorigenic role for PGE2, and suggest induction of 15-PGDH as potential way for the prevention and treatment of human cholangiocarcinoma.