AFP Knockout Huh-7 Cell Line

The AFP Knockout Huh-7 Cell Line is a CRISPR/Cas9-edited knockout cell line in which the human alpha-fetoprotein (AFP) gene has been disrupted. This loss-of-function model is derived from the Huh-7 hepatocellular carcinoma cell line, providing a genetically stable system for investigating AFP-dependent processes.

The Huh-7 cell line was isolated from a liver tumor of a 57-year-old Japanese male and serves as a well-characterized hepatic epithelial model. These cells are widely utilized in hepatology research, particularly in studies of hepatitis C virus replication, hepatocellular carcinoma biology, and hepatocyte signaling pathways, owing to their retention of hepatocyte-specific functions.

AFP is a fetal serum glycoprotein that binds fatty acids, bilirubin, and copper, and modulates proliferation, apoptosis, and immune evasion. Transcriptionally regulated by HNF1??, HNF4??, C/EBP??, p53, TGF-??, and retinoic acid, AFP activates downstream PI3K/Akt and ERK1/2 (MAPK1/3) signaling to promote cell survival and proliferation. It also engages the Wnt/??-catenin pathway via ??-catenin (CTNNB1) and GSK-3??, and inhibits apoptosis through Bcl-2 and Bax modulation. AFP interacts with the AFP receptor (AFPR) to mediate retinoic acid transport and immune modulation, including inhibition of dendritic cell maturation. CRISPR/Cas9-mediated knockout of AFP in Huh-7 cells abrogates these oncogenic functions, reducing PI3K/Akt and ERK pathway activation and enhancing apoptosis. The disruption of AFP-AFPR interaction further impairs retinoic acid signaling and immune escape mechanisms.

Since Huh-7 cells natively express AFP, the knockout line provides an isogenic tool for dissecting AFP’s role in hepatocellular carcinoma. It enables precise analysis of AFP-dependent tumorigenic properties, drug sensitivity, and metastatic potential. As a diagnostic and prognostic biomarker, AFP validation is critical, and this model aids in evaluating compensatory signaling and therapeutic targeting strategies in liver cancer.

Applications include mechanistic studies, drug screening, tumor immunology, and biomarker validation. Researchers can utilize assays such as western blotting for AFP and phosphorylated signaling proteins, cell proliferation and apoptosis assays, soft agar colony formation, migration/invasion assays, tumor xenografts, and RNA-seq. The cell line is also suitable for liver regeneration, differentiation, and virus-host interaction studies. For further information, contact Ascent Research.