Effect Of Shikonin On The Function Of Gap Junction Composed Of Connexin32

Objective: To investigate the effect of shikonin in different concentrations on gap junction intercellular communication and the expression of Cx32 protein in Cx32-transfected Hela cells,exploring the mechanism.Methods: Quantitative Real-time PCR and Western-blot was used to verify the transfection and stable expression of Cx32-transfected Hela cells.To find the concentration of the shikonin used for GJ was detect by“CCK-8 assay”.The function of GJIC was determined by “Parachute”dye-coupling assay;Western-blot was performed to detect the expression level of Cx32.Standard colonyforming assay was used to determine effect of shikonin on cisplatin cytotoxicity.Results: Cx32 amplification in the Hela cells induced with do xycycline has the expression of Cx32,for no doxycycline induced Hela cells no Cx32 amplification,no expression of Cx32.For no doxycycline induced Hela cells,shikonin had no cytotoxicity on cells at the concentration of 0-4?M,for Hela cells induced with doxycycline,Shikonin had no cytotoxicity on cells at concentration of 0~3?M;Preatreatment of Hela cells with shikonin(1-3?M)could increased the intercellular dye transfer and the Cx32 protein expression.Treatment of cells with3?M shikonin increased the cisplatin toxicity only in the presence of functional GJ.Conclusion: Verify the successful transfection of Cx32 plasmid,and constructed a cell line which can regulate the expression of Cx32.Shikonin could enhance the function of gap junction in Hela cells,through increasing the expression of Cx32.Shikonin can enhanced cisplatin cytotoxicity,and has a sensitizing effect.