p21 transduction augments differentiated phenotypes in immortalized human hepatocytes *
Recently, there has been great expectation regarding the use of cell therapies such as hepatocyte transplantation and bioartificial livers to treat patients with liver failure. Improvement of differentiated cellular functions is of fundamental importance in hepatocyte-based biological therapies. The molecule of p21, known as WAF1, is a potent cyclin-dependent kinase inhibitor which regulates the transition from the G1 phase to the S phase in a cell cycle. Investigators have demonstrated that p21 transduction induces cellular differentiation in various cell lines. We used SV40Tag-immortalized human NKNT-3 hepatocytes in this study. A replication-deficient adenovirus vector, Ad5CMVp21, expressing a p21 gene under the control of the CMV promoter, was used to achieve efficient p21 delivery. A pTAT-p21 fusion protein was also utilized for transduction in NKNT-3 cells. Morphological alteration, cell-cycle progression, and protein expression of albumin, CYP3A4, and CYP2C9 were analyzed in the p21-transduced NKNT-3 cells. Immunofluorescent staining and Western blotting analysis for p21 showed efficient p21 transduction in the NKNT-3 cells using Ad5CMVp21 or pTAT-p21. Under cell cycle analysis, transduction of p21 caused G1-arrest in NKNT-3 cells, leading to differentiated hepatic phenotypes including a decreased N-C (nucleo-cytoplasmic) ratio, decreased cell density, and enhanced protein expression of albumin, CYP3A4, and CYP2C9. In the results presented here, we show that exogenous expression of p21 augmented cellular differentiation in immortalized human NKNT-3 cells. (Accepted on August 28, 2002) Kawasaki Igakkaishi 28(3) : 165-174, 2002
