Antiproliferative Effect Of Dracorhodin Perchlorate On Rabbit Keratocyte In Vitro

Objective:To investigate the antiproliferative effect of dracorhodin perchlorate on rabbit keratocyte in vitro.Methods:Rabbit keratocyte was cultured by tissue culture.Cultured cells were identified by immunocytochemistry staining using anti -vimentin monoclonal antibody and cytokeratin 12 goat polyclonal antibody.Growth characteristic of cells was observed by inverted phase contrast microscope in vitro,growth curve was drawing by MTT colorimetry ,population doubling time was calculated by counting the cell number.MTT colorimetry was used for assaying various concentrations of dracorhodin perchlorate on the inhibitory rate of cell growth within 24h,48h,72h, inhibition rate and IC50 were calculated.Inverted phase contrast microscope and electron microscope were used to observe the effect of dracorhodin perchlorate on morphology of rabbit keratocyte within 24h.Flow cytometry was used to analyze the distribution of cell cycle and apoptosis within 24h.Results:Rabbit keratocyte was cultured by tissue culture after 8-10d. Culured cells were positive for anti-vimentin immunofluorescence stain. Cell growth curve was approximate s-shaped, population doubling time was (30.73±2.47)h.MTT colorimetry showed that with the increasing of dracorhodin perchlorate concentration,A values taked on downtrend (P<0.001),inhibitory rate showed obviously docs-effect and time-effect dependence within 24h,48h,72h,and the IC50 of three times were 47.72μg/ml, 41.40μg/ml, 32.01μg/ml. Inverted phase contrast microscope and electron microscope showed that with the increasing of dracorhodin perchlorate concentration,the proportion of cell apoptosis grew.flow cytometry showed that with the increasing of dracorhodin perchlorate concentration,the G0/G1 cell proportion increased by degree (P<0.001),the cell proliferation index decreased by degree(P<0.001),cell apoptosis rate raised(P<0.001).Conclusion:Dracorhodin perchlorate had the effect of inhibiting the proliferation of rabbit keratocyte,and with the increasing of dracorhodin perchlorate concentration,cell apoptosis rate raised.