Effect And Mechanisms Of Icaritin On Saos2Cells

Objective To observe the effect of icaritin on osteosarcoma cell line Saos2andinvestigate the reasons of the inhibitory effects of icaritin and its induction ofapoptosis on Saos2cells.Methods Treat the Saos2cells with different concentration gradient drug;CCK8method to test the effects of Proliferation inhibition; use the Inverted phase contrastmicroscope to observe morphological changes of Saos2; AnnexinV/PI double markermethod to detect the early Saos2cells apoptosis induced by icaritin by using flowcytometry; Treat the Saos2cells and the pretreated cells with antioxidant withdifferent concentration gradient icaritin,and then observe the cell morphology withthe inverted microscope; Flow cytometry instrument detect the ROS and apoptosisratio of the cells under different condition; The scratch test detect its influence onmigration and proliferation of Saos2; RT-PCR detect the HIF1alpha, Caspase3, Bcl–2gene expression leval; use Chloride cobalt (100μM) to analog anoxic environmentof osteosarcomat,then RT-PCR to measure VEGF、uPAR、MMP-2、ADM、Enolase-1、aldolase A gene expression in cells.Results Icaritin can inhibit the proliferation of Saos2,and also can induceSaos2cells apoptosis，icaritin can increase the ROS of Saos2compare with thecontrol group, antioxidant can alleviate this effect and reduce the apoptosis ratio atthe same time. icaritin can down-regulate HIF-1α、Bcl-2and upregulate theCaspase-3gene expression level, icaritin can also reduce the VEGF、MMP2、ADM、Enolase-1gene expression.Conclusions Icaritin can inhibit Soas2cells’ proliferation and induce the earlyapoptosis, proliferation inhibition may be associated with down-regulation ofHIF-1αand its downstream genes such as VEGF, MMP2、 ADM, Enolase-1expression.and up-regulation Caspase-3、decreased Bcl-2gene expression level andupregulation of intracellular reactive oxygen may take responsibility for theapoptosis.