Clear Cell Foci as Precursors of Hepatocyte Nuclear Factor 1-alpha–inactivated Hepatocellular Adenoma in a Metabolic Syndrome Mouse Model

Clear cell foci (CCF) are frequently observed in metabolic dysfunction-associated steatohepatitis (MASH) and are considered potential precursor lesions of hepatocyte nuclear factor 1α-inactivated hepatocellular adenoma (H-HCA). To clarify their chronological development, we examined 55 male TSOD mice at 24, 32, 40, and 48 weeks of age using histology and immunohistochemistry for glutamine synthetase (GS), liver fatty acid-binding protein (L-FABP), β-Klotho, and fibroblast growth factor 21 (FGF21). CCF first appeared at 24 weeks and increased markedly with age (from 11% to 81%). All CCF were positive for β-Klotho, and a subset showed FGF21 expression, indicating that CCF represent a hepatocellular state associated with metabolic dysregulation. H-HCA, characterized by GS negativity and reduced L-FABP expression, emerged at 40 weeks and reached an incidence of 29% at 48 weeks. Notably, multiple H-HCA were partially or completely surrounded by β-Klotho-positive CCF, suggesting a morphologic continuum from CCF to H-HCA. Raman spectroscopic analysis demonstrated that CCF exhibit prominent autofluorescence and possess spectral characteristics distinct from both background hepatocytes and tumor tissue, supporting the concept that CCF represent a unique hepatocellular state. These findings indicate that metabolic abnormalities in TSOD mice promote the sequential formation of CCF and H-HCA, establishing this model as a useful platform for studying adenoma development in metabolic liver disease.