Background and aim: PPARgamma is a nuclear receptor that regulates genes involved in energy metabolism. It consists of a transactivation domain at the N-terminus, two zinc fingers required for DNA binding, and a ligand-binding domain at the C-terminus that facilitates RXR-alpha binding and activation. The interaction of PPARgamma/beta-catenin has recently been established in type 2 diabetes and the development of colon cancer. On the other hand, Pirfenidone (PFD) has shown antifibrotic, anti-inflammatory, and antioxidant effects in various models of liver damage. The objective of our work was to demonstrate by in silico analysis that PFD is a ligand/agonist of PPARgamma and subsequently analyze the activity of beta-Catenin in the HepG2 hepatocarcinoma cell line.
Material and methods: Molecular interaction analysis was performed using the SwissDock platform, the images were made with the 3D UCSF CHIMERA processor. For in vitro analysis, the HepG2 cell line was used. The cells were treated with 500μM PFD, the non-selective agonist (GW7647; 100nM) and the selective antagonist (GW9662; 100nM) of PPARgamma for 24hrs. Immunofluorescence and Western-Blot of PPAR gamma and beta-Catenin were performed. The experiments were carried out in triplicate, GraphPadPrism 7 was used to prepare the graphs and statistical analysis.
Results:In silico analysis shows that Pirfenidone binds to the Serine342 residue of PPARgamma, the same site that Rosiglitazone binds to. Immunofluorescence shows increased PPARgamma placement and lower beta-Catenin in the nucleus for cells treated with PFD and GW7647. The opposite is observed in control and GW9662-treated cells. There is a differential expression of PPARgamma and beta-Catenin in cells treated with PFD and GW7647.
Conclusions: PFD is a ligand /agonist of PPARgamma because it binds to the Serine342 residue, just as Rosiglitazone does (a pharmacological agonist used in the treatment of type 2 diabetes mellitus). Additionally, treatment with PFD in HepG2 cells decreases the translocation of beta-Catenin to the nucleus, which could contribute to slow the progression of HCC.
This work has been partially subsidized by CONACyT basic science 259096 CB-2015-01. Asignated to JAB.
Conflicts of interest: The authors have no conflicts of interest to declare.
