The Protective Effects And Potential Mechanisms Of Kudiezi Injection On No-Reflow After Myocardial Ischemia-Reperfusion In Rats

KuDieZi Injection?KDZI?,also called DieMaiLing Injection,is a purified Chinese traditional medicine injection,which is extracted from the whole plant of Ixeris Sonchifolia?Bge?Hance.The main active components are adenosine and flavonoids.During clinical treatment,it's applied to cardiovascular diseases,which possesses the ability to relax vessels,inhibit the aggregation of platelets.However,its effects and possible mechanisms on no-reflow?NR?remain unknown.Objective:Our aim is to find out whether KDZI possesses the capability to protect myocardial from experimental ischemia/reperfusion related no-reflow in rats and reveal the underlying mechanisms.Methods:136 Wistar?female?rats were divided into four groups randomly according to their body weight.The reperfusion-related NR model was induced by ligation of coronary LAD for2 hours,then release the ligation for another 2 hours as reperfusion.The administrations were given to different groups 3 days ahead respectively,once a day,in which rats of Sham and Model group were given 5 mL/kg Normal Saline by caudal vein injection and rats of KDZI low,high dose group were given KDZI for 5 mL/kg and 10 mL/kg by caudal vein injection,respectively.Surgery was given at day 3 right after administration.After 2-hour reperfusion,rat was anaesthetized,then Evans Blue,TTC,Thioflavin S staining were adopted to assess the AAR,AAI,AAN of myocardial and cardiac function was measured by Color Doppler ultrasound.In addition,ELISA and biochemical kit were utilized to gauge the activity of CK-MB,LDH,AST,SOD,MPO and the content of Hs-CRP,PGI₂,NO,TXA₂,ET-1,TNF-?,IL-17,IL-6 in serum.The histology changes were indicated by HE staining of rats'heart.The blood samples were adopted to detect Blood Viscosity,PAR,PAG?1,3,5 min?.Apart from that,the expression level of ICAM-1 and P-Selectin in heart tissues were examined by immunohistochemical staining.On top of that,the phosphorylation pattern of PI3K/Akt/eNOS in heart were calculated by Western Blot.Results:?1?Compared to Sham group,AAR,AAI and AAN of model group were significantly higher?P<0.001?.While compared with Model group,KDZI low,high group markedly decreased AAR and AAI?P<0.05 or P<0.001?,only KDZI high group showed a significant decrease in AAN?P<0.01?.?2?Pretreatment of KDZI both low and high dose mitigated myocardial dysfunction,which was justified by elevated FS and significant increase of EF while decreased LVIDs and LVIDd compared to Model group?P<0.05?.?3?Compared with Model group,Pretreatment of KDZI both low and high dose lightened the leakage of CK-MB from myocardial?P<0.05 or P<0.01?;KDZI high group decreased the leakage of LDH,AST?P<0.05?.?4?Compared to Sham group,there were evident red blood cell leakage and neutrophil infiltration in Model group.The extent of myocardial damage was alleviated by pretreatment of KDZI.?5?The blood tended to be more viscous in rats going through surgery.While the increase of blood viscosity was overtly reversed by the administration of KDZI in high?120/Sec?,low?20/Sec?shear forces?P<0.05 or P<0.01?;KDZI high group significantly reverse the blood viscosity in middle shear force?P<0.01?.These results indicated that the viscosity of blood was decreased by KDZI,exhibiting an improved hemodynamics.?6?Pre-administration of KDZI inhibited the increase of PAR and PAG?1,3,5 min?compared with Model group?P<0.05 or P<0.01?.This results proposed that activation of platelets was inhibited by KDZI,resulting in the inhibitory effects on platelets aggregation and thrombogenesis.?7?After received KDZI,the decline of PGI₂ and NO,which both possess the ability to relax cardiovascular,were abolished compared with Model group.However,the increase of TXA₂and ET-1,which both are factors capable of contracting the cardiovascular,were abrogated compared with Model group?P<0.05,P<0.01 or P<0.001?.It's evident that normal function of vascular was preserved by KDZI.?8?It's also showed that pretreatment of KDZI reverted the decrease of activity of SOD and the increase of content of MDA in Model group?P<0.01 or P<0.001?.There was evident increase in the contents of Hs-CRP,TNF-?,IL-17,IL-6 and the activity of MPO in serum of Model group?P<0.001?.While pretreatment of KDZI significantly converted that trend?P<0.05,P<0.01 or P<0.001?.With these performance,it's clear that KDZI possess the capability to enhance the anti-oxidant system,down-regulate the activation of neutrophils,decrease inflammation in rats.?9?The positive staining of ICAM-1 and P-Selectin in heart tissues of Model group were markedly increased compared with Sham group?P<0.001?.While pretreatment of KDZI significantly ameliorated the expression of ICAM-1 and P-Selectin compared with Model group,which was dose-dependent?P<0.01 or P<0.001?.It's suggested that activation of both endothelial cells and platelets were inhibited by KDZI,followed by decrease in adhesion of endothelium-neutrophils and�platelets,thus resulting in suppressed thrombus formation and neutrophils infiltration.?10?The inhibition of phosphorylation of PI3K/Akt/eNOS in Model group were obviously abrogated by pretreatment of KDZI?P<0.01 or P<0.001?,which was also dose-dependent,suggesting the activation of PI3K/Akt/eNOS pathway induced by KDZI.Thus,NO was produced,exhibiting an effects of relaxation to blood vessel.Conclusion:In our study,it was proved that KDZI could prevent myocardial from ischemia/reperfusion related NR.The underlying mechanisms through which KDZI exerts its capabilities might be related with upregulation of anti-oxidant system,inhibition of inflammation and microthrombus formation,preserved normal function of vascular.