Hepatic stellate cells are considered to be liver‐specific pericytes that play a key role in liver fibrosis. Because these cells express desmin and smooth muscle α‐actin, they were assumed to be of myogenic origin. This hypothesis became doubtful when it was reported that stellate cells also express glial fibrillary acidic protein and neural cell adhesion molecule. In the present study, we show that activated stellate cells express nestin, a class VI intermediate filament protein originally identified as a marker for neural stem cells. Expression of nestin was first studied during spontaneous activation of stellate cells in culture. Immunohistochemistry showed that nestin‐positive stellate cells already appeared at day 3, and nearly all the cells became positive for nestin at day 6 and 15. The immunoreaction was present in filaments except in dividing cells. The presence of messenger RNA transcript for nestin was shown by reverse transcription polymerase chain reaction and sequencing of amplified complementary DNA. We then compared the presence of nestin with that of other intermediate filament proteins and smooth muscle α‐actin. Immunoblotting showed that the relative concentrations of nestin, desmin, and vimentin increased between day 2 and 6 in primary culture. After the initial increase vimentin leveled off, while nestin and desmin showed a tendency to decrease. This pattern was quite different from that of glial fibrillary acidic protein, which kept declining, and smooth muscle α‐actin, which increased continuously up to day 13 in culture. We then studied the presence of nestin in normal and CCl₄‐injured rat liver. In normal liver, minimal immunoreaction for nestin was observed within the liver parenchyma. During induction of fibrosis by carbon tetrachloride, nestin‐positive stellate cells appeared at 6 weeks, which was late in comparison with the induction of desmin and smooth muscle α‐actin. We conclude that nestin is induced in stellate cells during transition from the quiescent to the activated phenotype; culture activation is a stronger stimulus than in vivo activation by injection of CCl₄. Taken together with reports on expression of glial fibrillary acidic protein and neural cell adhesion molecule by stellate cells, new experimental studies on the embryonic origin of these cells are required