Hepatic management of toxic sterols after acute deletion of Cyp51 from cholesterol synthesis

Lanosterol 14α-demethylase (CYP51) is an enzyme involved in cholesterol synthesis, crucial for the normal liver function. Diminished activity of CYP51 leads to metabolism associated liver disease, ending in hepatocellular carcinoma. It is still not clear which processes are most affected in the hepatocytes and how they communicate with other cells towards the progressive liver pathology. Herein we describe a new inducible, liver-specific Cyp51 knockout mouse model (iLKO), developed to study how acute disruption of cholesterol synthesis is managed in the adult liver. Doxycycline inducible deletion avoided developmental confounders of albumin-Cre models and enabled isolation of viable primary hepatocytes. iLKO hepatocytes and liver tissue showed CYP51 depletion with accumulation of lanosterol and 24,25-dihydrolanosterol, while hepatic cholesterol levels remained largely unchanged, indicating compensatory uptake and/or pathway rerouting. Histological examination and transmission electron microscopy (TEM) revealed hepatomegaly with mild portal inflammation and ductular reaction but no overt fibrosis at the studied time points. iLKO hepatocytes displayed increased nuclear lipid droplets (LD) that might be involved in adaptation to endoplasmic reticulum (ER) stress. Additionally, we discovered crystal-like inclusions, especially in Kupffer cells. MALDI-TOF MSI could not resolve their composition, but their occurrence alongside sterol overload implicates non-cholesterol sterol crystallization as a potential trigger of inflammation. In summary, iLKO is a suitable model for dissection of sterol toxicity, clearly separated from developmental effects of Cyp51 depletion. Future examinations could reveal how toxic sterol intermediates are buffered in adult liver and how they might be connected to inflammation driven pathologies, relevant also to human health.