〔Abstract〕 Objective To investigate the effects of RO or incubation with cholesterol(CH) on the differentiation and apoptosis of KCs. Methods Ca2+(1.8 mmol/L) was added to KCs for 1 day after co-culture of RO alone or combined with CH in KCs for different time. The expression of Involucrin(INV) and Loricrin in KCs was analyzed by Western blot. After co-culture of RO alone or combination with CH in KCs for different time, the apoptotic changes of KCs cells were detected by flow cytometry and the expression of apoptotic proteins Bax and Bcl-2 were verified by Western blot.Results RO down-regulated the expression of Ca2+induced differentiation marker protein INV, but weakly inhibited the expression of Loricrin, while CH showed no antagonistic effect on RO. RO induced apoptosis of KCs cells in a time dependent manner. CH could antagonize the apoptotic effect of RO on KCs; the expression of Bcl-2 and Bax was inhibited when RO was applied alone,CH could partially antagonize the inhibition effect of RO on Bcl-2 expression, but had no significant effect on Bax; however, RO reduced the ratio of Bcl-2/Bax in a time-dependent manner, and CH partially weakened the effect of RO on the ratio of Bcl-2/Bax.Conclusion RO may inhibit KCs differentiation and induce KCs cell apoptosis by down-regulating the expression of Loricrin and Bcl-2.