Hemopexin Suppresses Hepatocellular Carcinoma via TNF-α-Mediated Mitochondrial Apoptosis

Fibrinolysis plays a crucial role in maintaining coagulation homeostasis, but its functions in hepatocellular carcinoma (HCC) remain poorly understood. This study aimed to develop a fibrinolysis-based molecular classification and prognostic signature for HCC and to identify a key regulatory gene. Using non-negative matrix factorization (NMF), we identified distinct fibrinolysis-related HCC subtypes with specific clinical outcomes and tumor microenvironment characteristics. A six-gene prognostic signature comprising ACAT1, GRHPR, HPX, PCK2, IYD, and PON1 was established through weighted gene co-expression network analysis (WGCNA) and LASSO-Cox regression, which effectively stratified patients into different risk groups across multiple cohorts. Hemopexin (HPX) was identified as the top candidate and functionally validated: HPX overexpression suppressed clonogenicity and migration, promoted apoptosis, and inhibited xenograft tumor growth. RNA sequencing analysis suggested associations between HPX and apoptosis as well as TNF-α signaling pathways, which were confirmed through flow cytometry apoptosis assays, mitochondrial membrane potential measurements, and TUNEL staining. Western blot and immunohistochemical analyses further demonstrated that HPX upregulates the Bax/Bcl-2 ratio via the TNF-α signaling pathway. This study defines novel molecular subtypes of HCC and reveals that HPX exerts anti-tumor effects through TNF-α-mediated mitochondrial apoptosis, characterized by an increased Bax/Bcl-2 ratio.