Z Gastroenterol 2019; 57(01): e43
DOI: 10.1055/s-0038-1677158
3. Metabolism (incl. NAFLD)
Georg Thieme Verlag KG Stuttgart · New York

Acetaminophen (APAP)-induces a dynamic response of the Hippo pathway and its effector yes-associated protein (YAP) in hepatocellular cells

L Aprupe
2   Modelling of Biological Processes, Centre for Organismal Studies/Bioquant, Heidelberg University, German
,
K Liu
1   University Hospital Heidelberg, Germany
,
J Schmitt
1   University Hospital Heidelberg, Germany
,
SE Weiler
1   University Hospital Heidelberg, Germany
,
A Ghallab
3   IfADo, Leibniz Research Centre for Working Environment and Human Factors at TU Dortmund, Germany
,
JG Hengstler
3   IfADo, Leibniz Research Centre for Working Environment and Human Factors at TU Dortmund, Germany
,
P Schirmacher
1   University Hospital Heidelberg, Germany
,
U Kummer
2   Modelling of Biological Processes, Centre for Organismal Studies/Bioquant, Heidelberg University, German
,
K Breuhahn
1   University Hospital Heidelberg, Germany
› Author Affiliations
Further Information

Publication History

Publication Date:
04 January 2019 (online)

 

Non-idiosyncratic drug-induced liver injury (DILI) after high-dose exposure to acetaminophen (APAP) is a very frequent cause for acute liver failure. APAP-induced hepatotoxicity is characterized by oxidative and endoplasmatic reticulum stress, sterile inflammation and microcirculatory dysfunction, as well as impaired hepatocellular regeneration. However, the connection between APAP-dependent DILI and the Hippo pathway, which is a key regulator of regenerative processes, inflammation, and the vascular network, is not understood.

We observed that the pro-proliferative Hippo pathway effector yes-associated protein (YAP), which represents a strong oncogene in the liver, is dynamically regulated in cancer cell lines upon APAP administration (5 – 10 mM; HLF and Hep3B cells). Between 1 – 4 hours a general YAP phosphorylation/inactivation was detectable, while at later timepoints (8 – 24 hours) YAP-dephosphorylation/activation followed by degradation after 24 – 48 hours was observed. Similar dynamic behavior was detectable for proteins of the central Hippo pathway kinase cassette (e.g. Lats1). As expected, several known YAP target genes such as CYR61, ANKRD1, CDC2 were induced at a time point where YAP is activated by APAP (24 hours). This induction was YAP dependent, since it was partly abrogated when YAP was silenced using gene-specific siRNAs.

To show the dynamic impact of APAP on YAP in normal hepatocytes, mouse livers were isolated after APAP injection (300 mg/kg; 1 hour to 16 days). Immunohistochemical analysis illustrated an oscillating enrichment of YAP nuclear with peaks at 12 hours and 4 days. Importantly, highest YAP target gene expression (mCYR61, mANKRD1, mCDC2) was detectable after 12 hours to 48 hours, which confirms the temporary and controlled regulation of YAP by APAP.

To understand how dynamically localized YAP may affect the biological outcome of Hippo/YAP signaling in hepatocellular cells, a minimal ordinary differential equation (ODE)-based computational model was created (incl. YAP, its paralogue TAZ, the upstream regulators Lats1/2, scaffolding factors such as 14 – 3-3 proteins, and 20 reactions). This mathematical model was fitted to spatial data derived from hepatocellular cells expressing RFP-labelled YAP and cultured under varying cell density conditions and will be used to describe APAP-mediated Hippo/YAP dynamics.

These data illustrate that APAP induces a highly dynamic response of YAP in hepatocellular cells (cancer cells and hepatocytes), which also affects expression of known YAP target genes. This process is not unspecific due to toxicity but depends on the activity of central upstream regulators of the Hippo kinase cassette. Further in vitro, in vivo, and in silico analyses will clarify if this specific process represents a cell-protective cellular response against APAP-mediated liver damage. Interestingly, these results would suggest that APAP may cause a time-limited cellular state with nuclear enrichment of the oncogene YAP, which is associated with elevated tumor risk.