| ObjectiveTo observe the changes of TGF-?1/Smads signaling on myocardial fibrosis cells induced by isoproterenol and clarify the effect of a s t r a g a l u s p o l y s a cch a r i d e on attenuating myocardial fibrosis cardiomyocytes and the underlying mechanism.MethodsIn the experimental of animals,fifty rats were divided into six groups by the double blind method : control group,ISO group,ISO+PRO group,ISO+ APS(200 mg?kg-1?d-1,i.g.)group,ISO+APS(400 mg?kg-1?d-1,i.g.)group and ISO+APS(800 mg?kg-1?d-1,i.g.)group.Rats received ISO,PRO or APS administration for 4 weeks by ig after intraperitoneal injection of ISO for 2 weeks.Sirius Red Staining were used to measure fibrotic degree of cardiomyocytes.Hemodynamic indexes were used as left ventricular function.Elisa was used to detect carboxyterminal propeptide of type I procollagen a n d type I collagen cross-linked C-terminal telopeptidei n r a t s s er um.Western blot was used to analyze proteins expression of T G F-?1,Smad2/3,Smad4 a n d Sm a d 7 i n t h e m y o c a r d i u m o f r a t s.Real-time PCR was used to analyze proteins expression of T G F-?1 m RN A.In the experimental study of cells,primary cultured rat cardiac fibroblast cells were randomly divided into six groups: control group,ISO group,ISO+PRO group,ISO+ APS(25 mg?L-1)group,ISO+APS(50 mg?L-1)group and ISO+APS(100 mg?L-1)group.PRO and APS were added to treatment for 48 hafter ISO treatment for 48 h.Fluorescent quantitation were used to detect the number of fibroblasts.Elisa was used to detect the volume of carboxyterminal propeptide of type I procollagen a n d type I collagen cross-linked C-terminal telopeptide i n fibroblasts.Western blot was used to analyze proteins expression of T G F-?1,Sm ad2/ 3,Sm ad4 and Smad7 in t h e fibroblasts.Real-time PCR was used to analyze proteins expression of T G F-?1 m R N A.ResultsCompared with control group,after the success of the isopropyl adrenaline induced myocardial fibrosis,HWI and LVWI were significantly increased while ± dp / dtm a x were significantly decreased in the ISO group of myocardial tissue of rats.Meanwhile,In the ISO group,the myocardial cells were observed under light microscope.It was found that the fibers of the cardiac muscle were arranged in turbulence and broken.The inflammatory cell infiltration and extensive fibrous tissue were found in the stroma,and the volume integral of collagen was also observed.The expression of T G F-?1 protein,Smad2/3 protein and Smad4 protein increased by 98.0% and 85.0%,and the expression of Smad7 protein was decreased by 70.0%,and the expression of T G F-?1 and T G F-?1 m RNA in myocardial tissue was increased by 266.0%.In animal experiment,staining of Sirius red showed that CVF increased by 93.5% in the ISO group,compared with the normal group;APS(APS800,mg/kg)and PRO group decreased by 45.0% and 56.6% in the CVF.The interstitial fibrosis and the morphology and arrangement of myocardial cells in ISO+PRO group and administration group were improved greatly.In the cell experiment,the rate of myocardial fibroblast proliferation was decreased by 49.0% in the APS group.In animals and cells experiments,the protein expression of TG F-?1 in A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group was decreased by 41.3%,61.2%,73.5%,respectively.In the expression of TGF-?1 m RNA decreased by 35.5%,61.9%,71.4%,respectively in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group.APS function has been verified in the cardiac fibroblasts induced by ISO,the same performance for the protein expression T G F-?1 activity decreased by 12.3%,20.0%,31.0% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group.T G F-?1 expression of m RNA decreased by 15.4%,41.6%,56.6% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group.PICP,ICTP and PICP/ICTP were measured by ELISA.Compared with group ISO,the PICP decreased by 24.5%,30.0% and 42.8% in A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group,and the ICTP decreased by 27.0%,26.5% and 17.3% in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group,and the PICP/ICTP decreased by 4.8%,8.4% and 21.0% in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group.In cell experiments,the PICP decreased by 11.5%,28.8% and 34.3% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group,and the ICTP decreased by 1.02%,11.7%,18.1% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group,and the PICP/ICTP decreased by 10.6%,19.3% and 19.6% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group.It showed that Astragalus polysaccharides can inhibit the interstitial fibrosis of myocardial tissue.Astragalus polysaccharides affect the expression of Smads protein through TGF-?1 protein,protein expression of Smad2/3 protein in animal level,decreased the protein expression 84.0%,34.3%,52.3% in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group,compared with the ISO group;Smad4 protein expression compared with ISO group,the protein level decreased 9.1%,21.2%,32.9% in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group.Compared with the ISO group,the expression of Smad7 protein increased by 22.4%,104.4%,157.6% in the A P S(200 mg?kg-1)group?APS(400 mg?kg-1)group?APS(800 mg?kg-1)group.Cell experiment,compared with the normal group,the protein expression of Smad2/3 in ISO group compared with A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group,decreased the protein expression of 39.0%,27.0%,46.7%;the protein expression of Smad4 compared with the ISO group,the protein level decreased 10.0%,25.2%,62.7% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group.Compared with the group ISO,the protein expression of Smad7 increased by 93.8%,142.8%,205.3% in the A P S(25 mg?L-1)group?APS(50 mg?L-1)group?APS(100 mg?L-1)group.ConclusionAPS could significantly attenuate myocardial fibrosis cells induced by ISO and the anti-apoptosis effect was related to TGF-?1/Smads signaling pathway. |
PDF Full Text Request