This study investigates the role of Semaphorin 3C (Sema3C) in the progression of hepatocellular carcinoma (HCC). Sema3C is significantly upregulated in fibrotic liver, HCC tissues, and peripheral blood of HCC patients, as well as in sorafenib-resistant tissues and cells. Overexpression of Sema3C correlates with stemness properties in HCC. The study identifies NRP1 and ITGB1 as key receptors of Sema3C, which activate the AKT/Gli1/c-Myc signaling pathway to enhance HCC self-renewal and tumor initiation. Sema3C also facilitates extracellular matrix (ECM) contraction and collagen deposition in vivo, promoting hepatic stellate cell (HSC) activation and transformation into cancer-associated fibroblasts (CAFs). Mechanistically, Sema3C interacts with NRP1 and ITGB1 in HSCs, activating downstream NF-κB signaling, which stimulates IL-6 production and cholesterol synthesis in HSCs. Additionally, CAF-secreted TGF-β1 activates AP1 signaling to upregulate Sema3C expression in HCC cells, creating a positive feedback loop that accelerates HCC progression. Blockade of Sema3C effectively inhibits tumor growth and sensitizes HCC cells to sorafenib in vivo. These findings highlight Sema3C as a novel biomarker that facilitates the crosstalk between cancer stem cells and the tumor stroma, offering a potential avenue for enhancing treatment efficacy and overcoming drug resistance in HCC.This study investigates the role of Semaphorin 3C (Sema3C) in the progression of hepatocellular carcinoma (HCC). Sema3C is significantly upregulated in fibrotic liver, HCC tissues, and peripheral blood of HCC patients, as well as in sorafenib-resistant tissues and cells. Overexpression of Sema3C correlates with stemness properties in HCC. The study identifies NRP1 and ITGB1 as key receptors of Sema3C, which activate the AKT/Gli1/c-Myc signaling pathway to enhance HCC self-renewal and tumor initiation. Sema3C also facilitates extracellular matrix (ECM) contraction and collagen deposition in vivo, promoting hepatic stellate cell (HSC) activation and transformation into cancer-associated fibroblasts (CAFs). Mechanistically, Sema3C interacts with NRP1 and ITGB1 in HSCs, activating downstream NF-κB signaling, which stimulates IL-6 production and cholesterol synthesis in HSCs. Additionally, CAF-secreted TGF-β1 activates AP1 signaling to upregulate Sema3C expression in HCC cells, creating a positive feedback loop that accelerates HCC progression. Blockade of Sema3C effectively inhibits tumor growth and sensitizes HCC cells to sorafenib in vivo. These findings highlight Sema3C as a novel biomarker that facilitates the crosstalk between cancer stem cells and the tumor stroma, offering a potential avenue for enhancing treatment efficacy and overcoming drug resistance in HCC.