A New Mechanism For Honokiol To Inhibit Ang ?-induced Cardiac Hypertrophy

Objectives To explore the effect of honokiol on AII-induced myocardial hypertrophy and its possible mechanism in SD rats.Methods The myocardial hypertrophy model in SD rats was established by subcutaneous implantation of AngII capsule osmotic pump(the fill volume was over 90% of the instruction).Animal tests: Thirty-six male SD rats were randomly divided into 4 groups: Control group: A? group [A?:200ng/(kg·min)],A?+2.5HKL[A?:200ng/(kg·min)+HKL:2.5mg/(kg·d)],A?+5HKL[A?:200ng/(kg·min)+HKL:5mg/(kg·d)];Blood pressure of rats was measured by tail-cuff method;The cardiac function was examined by color doppler ultrasound;The level of A?,ANP and BNP were determined by radio immunoassay;HE and SR staining to observe myocardial morphological changes and myocardial collagen deposition;Western blot(WB)was used to detect the protein levels of TR3,P70S6 K,LKB1,etc.Cell tests: The protein levels of TR3,P70S6 K,LKB1 and other protein in cardiomyocytes were detected by Western blot;The mRNA levels of TR3 and LKB1 were detected by qPCR;Transfection expertment was used to detect the effect of overexpressed TR3 on LKB1/P-LKB1;Localization of TR3 and LKB1 in cells were observed by confocal laser microscopy;Detection of the expression of TR3 and LKB1 in cytoplasm and nuclear by nuclear cytoplasmic separation kit.ResultsAnimal tests:(1)Compared with the control group,the blood pressure of rats in AII group increased significantly;There was no significant difference in the blood pressure of rats in the AII group,AII+2.5HKL group and AII+5HKL group;(2)compared with the control group,the LAD,IVSD and LVPWD of rats in AII group increased significantly(P < 0.01);Compared with AII group,HKL decreased LAD,IVSD and LVPWD in a dose-dependent manner(P < 0.01);(3)Compared with control group,the level of serum AII in AII group was significantly increased(P < 0.05);There was no significant difference in AII level among AII group,AII+2.5HKL group and AII+5HKL group(P > 0.05);There was no significant difference in ANP and BNP level between four groups;(4)Compared with control group,the endocardium of AII group was thickened,the cross-sectional area of myocardial cells was increased,and the myocardial area was also increased,the levels of TR3,P-P70S6 K were significantly increased while LKB1/P-LKB1,P-AMPK were significantly decreased.HKL could reduce endocardium thickness,cross-sectional area of cardiomyocytes,collagen deposition,the levels of TR3,P-P70S6 K while increase the levels of LKB1/P-LKB1,P-AMPK in a dose-dependent manner.Cell tests:(1)Compared with the control group,levels of TR3,P-P70S6 K,ANP were increased significantly while the levels of LKB1/P-LKB1,P-AMPK were decreased significantly after treated with AII.HKL could inhibit the changes of these molecules induced by AII in a concentration dependent manner;.(2)Compared with the control group,the mRNA level of TR3 was up-regulated and the mRNA level of LKB1 was down-regulated after treated with AII,HKL could inhibit the mRNA changes of TR3 and LKB1 induced by AII;(3)Compared with the control group,when TR3 was overexpressed,the levels of LKB1/P-LKB1,P-AMPK were decreased significantly,and the changes of three molecules were reversed after treated with HKL;(4)Fluorescence experiments showed that TR3 and LKB1 were mainly located in the nucleus.Compared with the control group,the localization of TR3 in nucleus and cytoplasm were increased after treated with AII,and the localization of LKB1 in the nucleus was increased while the location of cytoplasmic was decreased.Compared with the AII group,the localization of TR3 in nucleus and cytoplasm were reduced with the treatment of HKL,and the localization of LKB1 in the nucleus was decreased while the location of cytoplasmic was increased.(5)Nuclear and cytoplasmic separation results showed that the expression of TR3 in nucleus and cytoplasmic were increased while the expression of LKB1 in cytoplasmic was decreased after treated with AII.And HKL could decrease the expression of TR3 and LKB1 in nucleus while increase the expression of LKB1 in cytoplasmic in a concentration dependent manner.Conclusions(1)AII promoted blood pressure increase,endocardium thickening,myocardial cell cross-sectional area increase,myocardial collagen deposition increase,up-regulated TR3,P-P70S6 K protein levels and down-regulated LKB1/P-LKB1 protein levels;(2)HKL can inhibit myocardial hypertrophy and levels of TR3,P-P70S6 K,LKB1/P-LKB1,P-AMPK induced by AII in a dose-dependent manner;(3)AII can promote TR3 synthesis at the transcription level and inhibit LKB1 synthesis;HKL can inhibit TR3 synthesis at the transcription level and promote LKB1 synthesis;(4)AII promoted the interaction of TR3 with LKB1,and LKB1 was more bound to the nucleus.HKL inhibited the interaction between TR3 and LKB1,which made LKB1 transfer from the nucleus to the cytoplasm,phosphorylated and activated its downstream AMPK molecule.