Myofibroblasts (MFs) as well as hepatic stellate cells (HSCs) are known to be involved in liver fibrogenesis. Quiescent HSCs (qHSCs) in culture have been thought to differentiate to replicative activated HSCs (aHSCs). In this study a qHSC-enriched fraction isolated by Nycodenz-isodensity centrifugation was separated with a fluorescence-activated cell sorter, which revealed the presence of a small fraction (occupancy rate=0.4%) of cells that did not show vitamin A-autofluorescence under ultraviolet (UV)-irradiation (UV- cells). The remaining vitamin A-containing cells were autofluorescent (UV+) and originally expressed markers of qHSCs, and, in culture, did not grow, lost vitamin A, and expressed markers of aHSCs. UV- cells showed morphology of MFs, and, in culture, grew to form colonies and expressed markers of MFs. These results indicated that UV+ and UV- cells represent qHSCs and MFs, respectively, and that aHSCs have no growth potential and are a cell-type distinct from proliferative MFs. Gene expression profiles of UV- cells (MFs) newly identified gremlin as one of MF-preferential genes and its proteins were localized around fibrotic septa in rat and human livers. In addition, we suggested that the qHSC-enriched fraction included approximately 6% of liver MFs.