Study On The Mechanisms Of Shikonin Induced Human Salivary Adenoid Cystic Carcinoma Cell Apoptosis In Vitro

ObjectiveTo investigate the effect of Shikonin on inhibiting proliferation and inducingapoptosis of human salivary adenoid cystic carcinoma cells and designed toprovide experimental evidence and theoretical basis for the clinical applicationof Shikonin anti-cancer drug as an adjunct to the treatment of adenoid cysticcarcinoma.Methods1. MTT assay was applied to determine the half inhibitory concentration ofeffects on human salivary adenoid cystic carcinoma SACC-83cells treatedwith Shikonin，after different concentrations of shikonin deal with humansalivary adenoid cystic carcinoma SACC-83cells24h,48h,72h.2. Morphological changes of apoptotic cells were observed underfluorescence microscope after30μmol/L shikonin deal with the humansalivary adenoid cystic carcinoma SACC-83cells24h after acridineorange/ethidium bromide staining.3. Flow cytometry with PI staining to detect cell cycle， after differentconcentrations of shikonin deal with human salivary adenoid cysticcarcinoma SACC-83cells24h.4. Expression of apoptosis-related protein caspase-9, caspase-8andcaspase-3were detected by the Western-Blot after differentconcentrations of shikonin deal with human salivary adenoid cysticcarcinoma SACC-83cells24h. Results1. MTT assay resules showed that: Shikonin on human salivary adenoidcystic carcinoma SACC-83cells proliferation inhibition its role was timeand does dependent (P＜0.05).2. The AO/EB fluorescence staining:30μmol/L Shikonin the role of humansalivary adenoid cystic carcinoma SACC-83cells24h under fluorescentmicroscope observation, the control group of normal living cells showed auniform green, normal structure; experimental group of SACC-83cellswas orange, was bead-shaped, membrane rupture, the typicalmorphology of apoptotic cells.3. Flow cytometry with PI staining results show: after24h by Shikonin role ofSACC-83cell apoptosis rate increased with the drug concentrationgradually increased S phase cells gradually increased,the percentage ofcells in G0/G1phase gradually decline, no significant changes in G2phasecells.4. Western-Blot results show that grass prime role of human salivary adenoidcystic carcinoma SACC-83cells24h, caspase-9、 caspase-8andcaspase-3expression increased, compared with the control group wassignificantly different (P＜0.05).Conclusions1. Shikonin have the effect of inhibiting proliferation and induces apoptosisfor human salivary adenoid cystic carcinoma SACC-83cells in vitro.2. Shikonin induced human salivary adenoid cystic carcinoma SACC-83cellsapoptosis mechanism may occur with cell cycle arrest in S phase is relatedto.3. The mechanism of inducing apoptosis for human salivary adenoid cysticcarcinoma SACC-83cells might be related by up-regulation expression ofcaspase-9,caspase-8and caspase-3.It might be induce apoptosis inhuman salivary adenoid cystic carcinoma SACC-83cells by theFas-dependent pathway and mitochondria pathway.