TGF-β2 in Chronic Liver Disease (CLD) and Hepatocellular Carcinoma (HCC)

TGF-β1 is a pro-fibrogenic cytokine in chronic liver diseases (CLD), driving fibrogenesis. Many experimental approaches to tackle fibrogenesis were directed towards TGF-β1 and provided promising results. However, none of these could be translated into clinical trials for patient treatment up to now. In contrast, clinical trials have been conducted that were directed against TGF-β2.

Participation of TGF-β2 in mechanisms of CLD progression and HCC was first determined on mRNA expression levels in comparison to TGF-β1. TGF-β2 is expressed in hepatic stellate cells (HSC) and hepatocytes (HC) from mouse. TGF-β2 expression levels increase with activation of HSC within a time course of 8 days. TGF-β1 stimulation could not induce TGF-β2 expression in HSCs. Further, TGF-β2 expression was examined in monolayer and sandwich cultures of HCs, representative for stages of cellular stress. It is much higher in the monolayer culture, suggesting its induction during cellular damage. On the protein level, TGF-β2 secretion of HSC and HCs were analysed and compared to TGF-β1. In both liver cell types, TGF-β2 secretion was present, however less than TGF-β1. To translate findings in mouse to human, we examined the human HSC line LX-2 and found out that TGF-β1 expression is 2-fold higher compared to TGF-β2, whereas protein secretion was equal. Next, we investigated TGF-β2 vs. TGF-β1 mRNA expression levels in 8 HCC cell lines and found higher TGF-β2 levels in 5 of these, whereas in the other three it was equal. Cell lines with higher TGF-β2 mRNA levels also secreted higher amounts of TGF-β2 protein. Further, we analysed 3 models of CLD (CCl4, BDL & Mdr2) and a liver regeneration model upon acute liver damage. In liver regeneration and fibrosis upon CCl4, HSC activation and collagen expression is accompanied with transient increases of TGF-β2 expression, whereby expression levels were at least equal for both TGF-β isoforms. BDL, a model for secondary biliary fibrosis, displayed a TGF-β2 elevation within a time course of 14 days that was even about 10-fold stronger compared to TGF-β1. Studies with sample cohorts of Mdr2-/- and TGFα-cMyc tg mice are ongoing. Even more importantly TGF-β2 expression was found significantly upregulated in 2 independent HCC patient collectives, one comprising 40 HCC patient (in reference to the mean value of 7 independent normal liver samples; Neumann et al., Hepatology 2012), and one with 27 HCC patients (referring to non-tumor tissue of the same patient (iCOD database) with an average fold change of ˜3. TGF-β1 was also found about 3-fold upregulated in the Neumann cohort, whereas it was found differently expressed and even down regulated in HCC patients from iCOD database.

Currently we investigate the downstream signaling pathway activated by TGF-β2 in HSC in comparison to TGF-β1. First approaches indicate that both, TGF-β1 and TGF-β2 can activate canonical Smad signaling, whereas the non-canonical pathway via MAPK-p38 kinase is only induced by TGF-β2. The more or at least equal induction of TGF-β2 besides TGF-β1 in CLD along with the availability of antisense oligonucleotides (AONs) that have already been approved for clinical use in patients, suggests a new TGF-β directed therapeutic approach to tackle liver fibrosis and HCC.