| ObjectiveFrom the perspective of molecular biology and cytokine network, the mechanism of the action on NF-κB signal transduction pathway may control the system of CHF,myocardial excessive activation on the correlation between the level of serum MIF,TNF-α,IL-6,TGF-β1; To verify Astragalus polysaccharide Tanshin-one IIA does excessive activaton on NF-κB signal transduction pathway.Point of view that Qi asthenia and stagnation of blood-immunoregulation, to explore the intervention effection on benefiting vital energy and activating blood circulation of traditional Chinese medicine; To explore the intervention and anti-inflammatory immune active ingredients, it may provide a new method to reduce cardiovascular hospitalization rates of CHF.Method1. To set up animal models of CHF:Used the coronary artery ligation method,decreased the feed a half after two weeks, swam after three weeks, and then were made ultrasound Doppler ECG after4weeks.2. The SD rats were randomly divided into six experiment groups:APS group(3g.kg-1.d-1,ig),Tanshinone group(5mg.kg-1.d-1,iv),APS+Tanshinone group(dose as above),Atorvastatin group(10mg.kg-1.d-1,ig),model group(the volume of normal saline,ig),sham group as negative control(to play virtual nodes ligation of coronary artery,then were given normal saline to the duodenum by dose method),medicines were given for6wk.3. Myocardial pathomorphology was observed by light microscope.4. Immunohistochemistry was used to detect the expression of NF-κB in myocardial tissues.5. The levels of NF-κB and MIFmRNA in the myocardial tissues were measured by RT-PCR and fluorescent real-time quantitative PCR method.6. Western blot was used to detect the gene expression of NF-κB,IκB and IkK,and software LabWorks was used to analyze the gray image data.7. the levels of MIF,TNF-α,IL-6and TGF-β1in the serum were measured and analyzed the correlation by ELISA.8. SPSS17.0statistical software was used to analyze data.Result1. Echocardiography results showed that model group and sham group were detected by cardiac ultrasound after4weeks, respectively, left ventricular end-diastolic diameter(LVEDD),left ventricular end diastolicvolume(LVEDV) of model group were higher than sham group,and ejection fraction(EF) of model group was lower than sham group(P<0.01).2. Pathological results showed that in model group there were many myocardial nuclear shrinking wall of local irregular, necrotic fociformation, decreased myocardial fibers,lighter myocardial stripes,and some necrosis replaced by collagenfiber formation of scar tissue,marked hyperplasia of connective tissues, large inflammatory cell infiltration; Remarks of medication groups wall damage decreased,increased residual myocardial cells,clearer stripes,visible scar tissue and inflammatory cells;Pathologic score of model group was higher increased than sham group(P<0.05);Pathologic scores of medication groups were significantly lower than those of model group(P<0.05); Remark of APS+Tan group was the lowest(P<0.05).3. Immunohistochemistry showed that the expression of NF-κB in model group was higher than sham group(P<0.05); that of medication groups were lower than that of model group(P<0.05);that of matching group was lower than that of single medication group(P<0.05).4. RT-PCR assay results showed that the expressions of NF-κB and MIFmRNA in model group were significantly higher than medication groups; medication groups had much more inhibition than sham group(P<0.05); those of matching group were lower than those of single medication group(P<0.05).5. Fluorescent real-time quantitative PCR results show that the target gene basis of NF-κB and MIFmRNA:sham group(1),(1); Atorvastatin group(1.239),(1.109); APS+Tan group(1.453),(1.515); APS group(1.777),(1.777); Tan group (1.986),(2.219); model group(2.345),(2.602); those of medication groups decreased significantly than sham group(P<0.05); those of APS+Tan matching group were lower than those of single medication group(P<0.05).6. Western blot showed that the expression of NF-κB in model group was significantly higher than sham group(P<0.05); that of medication groups was lower than model group(P<0.05); that of matching group was lower than single medication group(P<0.05); but on the contrary,the expressions of IκB、IκK in model group were lower than sham group(P<0.05).7. The levels of MIF,TNF-α,IL-6in the serum by ELISA showed that those of model group were(5.94±2.14),(420.36±67.57),(3704.44±1048.81),which were higher than sham group(0.23±0.08),(53.96±3.18),(352.40±87.20)(P<0.05); those of medication groups were significantly lower than those of model group(P<0.05). but on the contrary,the level of serum TGF-β1showed an opposite tendency. 8. Analysis of correlation between serum MIF、TNF-α、IL-6by ELISA showed that those of medication groups decreased significantly,and the level of serum MIF was positively correlated with those of serum TNF-α and IL-6(r1=0.879,r2=0.904, P<0.05),but the level of serum MIF was negative correlated with that of TGF-β1(r3=-0.602,P<0.05).Conclusions1. NF-κB as a hub in the signal transduction pathway released into the nucleus after activation,and accompanied by inhibition of IκB and degradation of most critical kinase IκK, all of which can suggest that NF-κB signal transduction pathway play an important role in CHF incidence.2. The levels of MIF in the serum and myocardial tissues of CHF rats significantly enhanced, and the expression of MIF, TNF-α and IL-6showed a positive correlation, but negatively correlation with those of serum TGF-β1, suggesting that as an upstream cytokine,MIF plays a key immunomodulatory role on onset of CHF.3. Astragalus polysaccharide joint Tanshinone-ⅡA immunomodulatory therapy can correct the immune status of CHF rats,regulating the equilibrium between inflammatory and anti-inflammatory cytokines,possessing significant anti-inflammatory and immunocompetence activity.Those of matching group were more effective than single medication group, blocking the overactivation of NF-κB transduction pathway,and MIF has shown a significant therapeutic curative effection.This study can provide a broaden prospect for anti-inflammatory immune and active ingredients of the traditional Chinese medicine in treating CHF. |
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