AIM: To show the efficient generation of hepatocytelike cells(HLCs) differentiated from the induced pluripotent stem cells(iP SCs) of rats.METHODS: Hepatic differentiation was achieved using a three-step protocol with several growth factors. First, rat i PSCs were differentiated into definitive endoderm cells using Activin A and Wnt3 a treatment. Then fibroblast growth factor 4 and bone morphogenetic protein 2 were added to the culture medium and used to induce hepatic differentiation. Finally, hepatocyte growth factor, Oncostatin M and dexamethasone were used for hepatic maturation. The liver-related markers and functions of HLCs were assessed at the gene and protein levels.RESULTS: After endodermal induction, the differentiated cells expressed endodermal markers forkhead box protein A2 and SRY-box containing gene 17 at the m RNA and protein levels. After 20 d of culture, the i PSCs were differentiated into HLCs. These differentiated cells expressed hepatic markers including α-fetoprotein, albumin CK8, CK18, CK19, and transcription factor HNF-4α. In addition, the cells expressed functional proteins such as α1-antitrypsin, cytochrome P450 1A2 and CYP 3A4. They acted like healthy hepatic cells, storing glycogen and taking up indocyanine green and low-density lipoproteins. Also, the rates of urea synthesis(20 d 1.202 ± 0.080 mg/dL vs 0 d 0.317 ± 0.021 mg/d L, P < 0.01) and albuminsecretion(20 d 1.601 ± 0.102 mg/d L vs 0 d 0.313 ± 0.015 mg/d L, P < 0.01) increased significantly as differentiation progressed.CONCLUSION: Rat i PSCs can differentiate into HLCs rapidly and efficiently. These differentiated cells may be an attractive resource for treatment of end-stage liver disease. AIM: To show the efficient generation of hepatocytelike cells (HLCs) differentiated from the induced pluripotent stem cells (iP SCs) of rats. METHODS: Hepatic differentiation was achieved using a three-step protocol with several growth factors. First, rat i PSCs were differentiated into definitive endoderm cells using Activin A and Wnt3 a treatment. Then fibroblast growth factor 4 and bone morphogenetic protein 2 were added to the culture medium and used to induce hepatic differentiation. Finally, hepatocyte growth factor, Oncostatin M and dexamethasone were used for hepatic maturation. The liver-related markers and functions of HLCs were assessed at the gene and protein levels .RESULTS: After endodermal induction, the differentiated cells expressed endodermal markers forkhead box protein A2 and SRY-box containing genes 17 at the m RNA and protein levels After 20 d of culture, the iPSCs were differentiated into HLCs. These differentiated cells express hepatic markers including α-fetopr They are characterized in that they express functional proteins such as α1-antitrypsin, cytochrome P450 1A2 and CYP 3A4. They acted like healthy hepatic cells, storing glycogen and taking up indocyanine Also, the rates of urea synthesis (20 d 1.202 ± 0.080 mg / dL vs. 0 d 0.317 ± 0.021 mg / d L, P <0.01) and albumin secretion (20 d 1.601 ± 0.102 mg / dL vs 0d 0.313 ± 0.015 mg / d L, P <0.01) increased significantly as differentiation progressed.CONCLUSION: Rat differentiated cells may differentiate into HLCs rapidly and efficiently. These differentiated cells may be an attractive resource for treatment of end-stage liver disease .