The lipogenic transcription factors Pparα and Pparγ as possible linkage between lipid metabolism and Hedgehog-Signalling Pathway?

As the biggest metabolic organ the liver performs a plethora of functions, including carbohydrate and lipid metabolism. The smallest functional unit thereby is displayed by the liver lobules showing a metabolic zonation. Hepatocytes can be roughly divided into periportal (PP) and pericentral (PC) cells. Numerous genes and enzymes are involved in the regulation processes of the metabolism; especially nuclear receptors like Ppars possess a coordinating role for many physiological functions.

While Pparα is assumed to promote fatty acid oxidation and, thus, is controlling lipid catabolism, Pparγ probably promotes the storage of lipids. Transcription factors like Ppars also interact with morphogenic pathways, such as the hedgehog (Hh) signalling pathway, which is crucial for embryogenesis and organogenesis. However, recently we hypothesized, that this pathway also plays a role in adult organs such as the liver, but molecular mechanisms have not yet been well characterized. Our aim was to analyze hepatocyte specific properties of the nuclear transcription factors Pparα and Pparγ in C57BL/6 mice with respect to their circadian rhythm and heterogeneous distribution within the parenchyma. Furthermore, we aimed to investigate the possible interactions between Ppars and Gli-transcription factors (TF) in adult hepatocytes.

To examine hepatic gene expression, RNA was extracted from whole liver lysates or hepatocytes. Additional in vitro knockdown experiments with siRNA against Pparα and Pparγ and a combination of both, as well as Gli-KD experiments, were carried out within a timeframe from 6 – 72 hours. Subsequently, in response to transfection, relative expression levels of target genes were determined by qRT-PCR. To further investigate the function of Pparγ, it was necessary to classify genes, which are influenced by siRNA-mediated KD against Pparγ. Alterations in gene expression of HH signalling components and lipid metabolism target genes were determined by using micro-arrays.

Generally our results showed that the nuclear receptors, Pparα and Pparγ, are heterogeneously distributed within the liver lobules. Both factors showed an altered expression due to their regulation by the circadian rhythm. KD-experiments of Ppars and Gli-TFs and their related mRNA expression profiles indicate a putative interaction between lipogenic TFs and Hh signalling. Pparγ-KD resulted in down-regulation of Gli1 and Gli3 after 48h, whereas Gli3-KD significantly up regulated both Ppar heterologues. Furthermore, the transcriptome analyses revealed that a wide variety of genes, like genes for biological processes (e.g. transports, cell cycle) are affected due to Pparγ-KD. These results allowed us to conclude that there is a mutual interaction between Pparγ and Gli3 at the transcriptional level. Future experiments should investigate whether these interactions are caused in a direct or indirect manner.