Effects Of Sodium Tanshinone ⅡA Sulfonate On Stably Expressed Human Cardiac I_Ks Current

BackgroundSlowly activating delayed rectifier potassium current(I_Ks) is important in therepolarization of human cardiomyocyte action potential,especially in forming andmaintaining plateau of action potential. I_Ks channel is combined byαandβsubunits,which are coded by KCNQ1 and KCNE1 respectively. Mutations ofhKCNQ1/hKCNE1 genes,chronic myocardial hypertrophy and chronic heartfailure will lead to dysfunction of I_Ks and ingenital or acquired LQTS which areimportant factor in hazardous ventricular arrhythmia such as Torsade depoints.Research into pharmacological properties of I_Ks channel may provide a newclue for protection and treatment of such arrhythmias.TanshinoneⅡA is a lipophilic form of salvia miltiorrhiza. SodiumtanshinoneⅡA sulfonate is soluble in water and is widely used to treat coronaryartery disease and myocardial infarction in clinic.However, Effects of sodium tanshinoneⅡA sulfonate on human cardiomyocyte slowly activating delayedrectifier potassium current are not known.ObjectiveObjectivesandMethodsPART I: Human cardiac KCNQ1 and KCNE1 genes, which encodeαandβsubunit of I_Ks respectively, were cotransfected into HEK 293 cells.Hygromycin Bwas used to screen the transfected cells.Electrophysiological and pharmacologicalmethods were used to identify whether properties of stably expressedhKCNQ1/hKCNE1 channels are similar to native human cardiac I_Ks channel.PARTⅡ:To investigate the effects of sodium tanshinoneⅡA sulfonate on I_Kscurrent by patch clamp. To make clear whether tanshinoneⅡA sulfonate can affectactivation curve of I_Ks current and activation and deactivation time constants ofI_Ks.ResultsPART I:1. Slowly activating current was recorded in HEK 293 cells stablely expressinghKCNQ1/hKCNE1 genes.The current was voltage-dependent obviously and itsactivation voltage was higher than ?40 mV. We could only record smallbackground current in HEK 293 cells without gene transfection.2．Its reverse potential linearly correlates with the extracellular potassiumconcentration,indicating that hKCNQ1/hKCNE1 current is a kind of potassiumcurrent.3．The current was significantly and reversibly inhibited by Chromanol 293B,aspecific I_Ks blocker, with an IC50 (+40mV)of 9.1μmol/L. 4．Extacellular fluid without potassium ions significantly increased the magnitudeof I_Ks. At + 40 mV, the increased level was (28.6±2.0)% (P＜0.01, n = 8).PARTⅡ:1．Sodium tanshinoneⅡA sulfonate reversibly enhanced I_Ks in a concentrationdependent way,EC50(＋40mV)is 64.5μmol/L.2．Sodium tanshinoneⅡA sulfonate at 30μmol/L produced a significant leftwardshift in the voltage-dependence of I_Ks activation without effect on slope factor. V0.5of sodium tanshinoneⅡA sulfonate at 30μmol/L and control groupwere(16.1±1.2)mV and(24.1±1.9)mV (P＜0.05,n= 7), S of these two groupswere 19.1±2.3 and 17.6±2.4 (P＞0.05,n=7).3．The activation time constants of I_Ks were reduced by tanshinoneⅡA sulfonate at-20 to +40mV(n=7,P＜0.05); however, the deactivation time constants wereincreased by sodium tanshinoneⅡA at potentials of -90 to -40 mV (n=7,P＜0.05).ConclusiConclusionPART I:Cell model stably expressing human cardiac slowly activating delayedrectifier potassium current has been successfully established, which has similarcharacteristics of native human cardiac I_Ks. The established cell model is a idealcell model of human cardiac I_Ks.PARTⅡ:Sodium tanshinoneⅡA sulfonate reversibly enhanced I_Ks in aconcentration dependent way. Sodium tanshinoneⅡA sulfonate at 30μmol/Lproduced a significant leftward shift in the voltage-dependence of I_Ks activationwithout effect on slope factor.Sodium tanshinoneⅡA sulfonate reduced activationtime constants of I_Ks ,but increased deactivation time constants of I_Ks.