The Role Of Shexiang Baoxin Pills And Ginsenoside Rb1 On Ventricular Remodeling After Myocardial Infarction Of Mice

Part ⅠThe effects of Shexiang Baoxin Pills and Ginsenoside Rb1 on ventricular remodeling and clinical outcomes after myocardial infarction of mice Objective: The adverse ventricular remodeling（PIVR）that occurs after myocardial infarction（MI）is an important factor leading to cardiac insufficiency and myocardial hypertrophy.The aim of this experiment was to investigate the effects of muscadine heart-preserving pill（SBP）and one of its main active ingredients,ginsenoside Rb1（GRb1）,on ventricular remodeling and survival rate and ventricular rupture rate after MI.Methods:C57BL/6 mice aged 7～9 weeks and weighing about 21 g were selected and randomly grouped,of which 1/4 of the mice were sham-operated group（Sham group）.The remaining were divided into three groups and established MI models,namely the SBP group,GRb1 group,and control group（Control group）.All four groups were fed under the same conditions,but the Sham group was not given any intervention,the SBP group was given 20 mg/kg SBP,the GRb1 group was given 20 mg/kg GRb1,and the Control group was given physiological saline.The administration was done by gavage once daily.The survival of the mice was recorded daily after MI,and the ventricular rupture was recorded by necropsy of the dead mice.Until day 21 post-MI,echocardiography was performed to detect cardiac function as well as ventricular chamber size in mice.Results:（1）The EF and FS values of mice in the Control,SBP and GRb1 groups were lower and the LVEDD,LVESD,LVEDV,LVESV values were higher than those in the Sham group on day 21 after MI.（2）The echocardiographic results showed that the EF and FS values of the mice in the SBP and GRb1 groups were higher and the LVEDD,LVESD,LVEDV,and LVESV values were lower than those in the Control group on day 21 after MI.（3）Mice in the SBP and GRb1 groups had a lower rate of heart rupture and a higher survival rate than the Control group on day 21 after MI.（4）No significant differences were found in EF,FS,LVEDD,LVESD,LVEDV,LVESV values as well as cardiac rupture rate and survival in the SBP group compared with the GRb1 group.Conclusion: SBP and GRb1 improved cardiac systolic function,reduced ventricular dilatation,improved PIVR,reduced ventricular rupture rate and increased survival in MI mice.Part Ⅱ The effect of Shexiang Baoxin Pills and Ginsenoside Rb1 on the polarization of monocytes / macrophages of miceObjective: SBP and GRb1 can regulate the secretory activity and polarization direction of immune cells,especially Mo / MФ.The Mo / MФ-mediated immune inflammatory response has been shown to play a key role in PIVR.The aim of this experiment was to investigate the effects of Shexiang Baoxin Pills（SBP）and Ginsenoside Rb1（GRb1）on the polarization of monocytes / macrophages after MI.Methods:C57BL/6 mice aged 7～9 weeks and weighing about 21 g were selected and randomly grouped,of which 1/4 of the mice were sham-operated group（Sham group）.The remaining were divided into three groups and established MI models,namely the SBP group,GRb1 group,and control group（Control group）.All four groups were fed under the same conditions,but the Sham group was not given any intervention,the SBP group was given 20 mg/kg SBP,the GRb1 group was given 20 mg/kg GRb1,and the Control group was given physiological saline.The administration was done by gavage once daily.After anesthetizing the mice on day 1,3,7 after MI,peripheral blood,bone marrow,spleen and heart infarct tissues were collected,and the ratio of Ly6C^high/Ly6 C^low monocytes in these tissues,as well as the ratio of Ly6C^high/M1 macrophages,Ly6C^low/M2 macrophages and neutrophils in the heart infarct area were measured by flow cytometry.The expression levels of IL-1β and chemokine MCP-1 in the peripheral blood and heart infarct area of MI mice were measured by ELISA and qRT-PCR,respectively.Results:（1）The proportion of Ly6 C^high monocytes in bone marrow,spleen,peripheral blood and heart infarct area tissues of mice in the SBP and GRb1 groups on days 3 and 7 after MI was lower than that in the Control group,and the proportion of Ly6 C^low monocytes was higher than that in the Control group.The proportion of Ly6 C^high monocytes was higher in the SBP group compared with the GRb1 group and lower in the Ly6 C^low monocytes compared with the GRb1 group in the bone marrow on day 3 after MI and in the spleen tissue on day 7 after MI,and no significant differences were found in the proportion of monocytes in each tissue compared with GRb1 at the remaining time points.（2）The proportion of Ly6 C^high / M1 macrophages in the tissues of the infarcted area of the heart on days 3 and 7 after MI in the SBP and GRb1 groups was lower and the proportion of Ly6 C^low / M2 macrophages was higher in the SBP and GRb1 groups than in the Control group.At day 3 after MI,the proportion of M1 macrophages was higher in the SBP group compared with the GRb1 group,and no significant differences were found in the proportion of macrophages at the remaining time points compared with the GRb1 group.（3）The levels of MCP-1 and IL-1β in peripheral blood and heart infarct area on days 3 and 7 after MI were lower in the SBP and GRb1 groups of mice than in the Control group.No significant differences were found in MCP-1 and IL-1β levels in peripheral blood and infarct area in the SBP group compared with the GRb1 group.（4）No significant difference was found in the neutrophil ratio in the infarcted area of the heart in the SBP and GRb1 groups of mice compared to the Control group.Conclusion: By observing the results,the changes in monocytes in peripheral tissues and in the infarcted area of the heart as well as macrophages in the infarcted area of the heart in the SBP and GRb1 groups of MI mice on the 3rd and 7th postoperative days responded to the promoting effect of SBP and GRb1 on the polarization of monocyte macrophages especially to the repair type(Ly6C^low / M2)after MI.In addition,the inhibitory effect of SBP and GRb1 on mononuclear macrophage production and chemotaxis was demonstrated by the observation of lower levels of MCP-1 and IL-1β in the peripheral blood and the infarcted area on postoperative days 3 and 7 in the SBP and GRb1 groups.Part Ⅲ The effect of Shexiang Baoxin Pills and Ginsenoside Rb1 on injury / repair after myocardial infarction by regulating the secretory function of macrophages of miceObjective: In previous studies,macrophages can regulate injury and repair after MI by secreting inflammatory factors with different effects（mainly embodied in: TNF-α,MMP-9,IL-6,IL-10,TGF-β,VEGF）at different stages of inflammation.The aim of this experiment was to investigate the changes in cytokine secretion by both types of macrophages and the eventual effects on injury and repair after regulation of macrophage polarity by Shexiang Baoxin Pills（SBP）and Ginsenoside Rb1（GRb1）.Methods:C57BL/6 mice aged 7～9 weeks and weighing about 21 g were selected and randomly grouped,of which 1/4 of the mice were sham-operated group（Sham group）.The remaining were divided into three groups and established MI models,namely SBP group,GRb1 group,and control group（Control group）.All four groups were fed under the same conditions,but the Sham group was not given any intervention,the SBP group was given 20 mg/kg SBP,the GRb1 group was given 20 mg/kg GRb1,and the Control group was given physiological saline.The administration was done by gavage once daily.The m RNA expression levels of Ly6 C^high / M1 macrophage-associated inflammatory factors TNF-α,MMP-9,IL-6 and Ly6 C^low / M2 macrophage-associated inflammatory factors IL-10,TGF-β,VEGF were measured by qRT-PCR on days 1,3,7 after MI.The infarct size,collagen deposition in the infarct area and fibrosis in the non-infarct area of each group of mice were detected using Masson staining on day 21 after MI.CD31 and α-SMA/Vimentin double staining,representing capillary and myofibroblasts,respectively,were detected using immunofluorescence staining,and type I collagen content was detected using Sirius red staining.Results:（1）The expression of inflammatory factors TNF-α,MMP-9 and IL-6 in the infarct area of MI mice in the SBP and GRb1 groups on postoperative day 3 and 7 was lower than that in the Control group,and the expression of IL-10,VEGF and TGF-β was higher than that in the Control group.（2）The infarct area of MI mice in the SBP group and GRb1 group was smaller than that in the Control group on postoperative day 21.（3）The capillary density of MI mice in SBP group and GRb1 group on the 21 st day after surgery was higher than Control group,and the proportion of myofibroblasts was higher than that in Control group.（4）The collagen content in the infarct area of MI mice in the SBP and GRb1 groups was higher on postoperative day 21 than that in the Control group,especially the type I collagen content was higher.（5）No significant differences were found in the expression of cytokines（TNF-α、MMP-9、IL-6,IL-10、VEGF、TGF-β）,infarct area,capillary density,myofibroblast ratio,and collagen deposition in the SBP group compared with the GRb1 group.Conclusion: SBP and GRb1 showed a decrease in the expression of Ly6 C^high / M1 MФ-related pro-inflammatory factors IL-6,TNF-α and MMP-9 and thus a decrease in infarct size,as well as an increase in the expression of Ly6 C^low / M2 MФ-related factors IL-10,TGF-β and VEGF after altering MФ polarization in MI mice,thus promoting myofibroblast activation,collagen,especially type I collagen.Fibroblast activation,collagen deposition,especially type I collagen,and capillary production.These changes ultimately affect the balance between injury and repair after MI,which in turn leads to improved PIVR.