The lysine specific histone demethylase LSD1 in activated Hepatic Stellate Cells contributes to liver fibrosis

Background and Aim:

Liver fibrosis is the ultimate progress of different kinds of chronic liver diseases, independently if chronic inflammation is caused by viral infection, alcohol abuse or other factors. Hepatic Stellate Cells (HSC), transdifferentiating into myofibroblasts during chronic inflammatory processes, are the central cell type of liver fibrosis. In response to myofibroblastic activation, HSC are characterized by high production of extracellular matrix proteins (ECM). Recent studies suggest that epigenetic mechanisms are important in triggering processes of liver fibrosis. The lysine specific histone 3 demethylase LSD1 is shown to be a central mediator of histone modification, which is crucial for tumor progression of different cancer types. However, its function in inflammatory and fibrotic processes is not yet known. Hereby, we aimed to study the function of LSD1 in cellular and molecular mechanisms of liver fibrosis.

Methods:

We inhibited LSD1 function in myofibroblastically activated human and murine HSC cells (LX-2, HSC-T6) using a LSD1 inhibitor compound. In addition, CRISPR/Cas mediated LSD1 knock-out HSC clones were generated. The influence of LSD1 silencing on profibrogenic HSC activation was further studied by expression profiling using hybridisation microarrays. Additionally, global protein expression and phosphorylation pattern was extensively analysed by protein mass spectrometry followed by signaling pathway analysis. The data was validated by immunoblotting and immunochemistry.

Results:

LSD1 inhibition led to a pronounced change in HSC morphology, viability and expression profiles. In response to LSD1 inhibition, HSC showed a morphology resembling quiescent HSC, and an altered expression profile of profibrogenic markers showing inhibition of collagen subunits, but increased synthesis of progenitor markers such as OCT-4. Further comprehensive transcriptomic and proteomic studies revealed that LSD1 mediates the control of downstream epigenetic targets, which are known to be related to liver fibrosis, such as histone deacetylases, histone methyl transferase and DNA-methyltransferases.

Conclusions:

Our studies demonstrate that LSD1 plays a central role in liver fibrosis by affecting multiple profibrotic epigenetic pathways.