| Cardiovascular disease is an important cause of human death, in which morbidityand mortality rates in the first place of ischemia heart diseases.In order to prevent theischemic myocardium from inevitable rescue necrosis, re-let the blood flow to theischemic myocardium is very important, generally percutaneous coronary intervention(PCI) or coronary artery bypass grafting (CABG) are used in clinical thrombolytictherapy. Unfortunately, thrombolytic therapy acuting myocardial ischemia can lead tomyocardial injury, described as "reperfusion injury"(I/R). As China entered the agingsociety, the incidence of myocardial infarction increased year by year, myocardialinfarction reperfusion injury has become the focus of attention in researchers, it hasimportant implications for studing myocardial ischemia and reperfusion injury indrugs.Pathogenesis of myocardial ischemia and reperfusion mainly has inflammatoryresponse, energy metabolism, oxygen free radicals outbreak, Ca2+overload andabnormal pH. JNK is the key target on inflammatory myocardial ischemia-reperfusioninjury, JNK can not only been activated in ischemia phase, ischemia/reperfusionphase has also been activated. This may also result in activation of the alreadyactivated NF-κB, translocation into the nucleus, involved in inflammation, and furthercause inflammatory injury. Therefore, JNK may play a key role in the regulation ofreperfusion injury. Studies have shown that JNK knockout mouse heart hadsignificantly reduced ischemia/reperfusion injury infarction and apoptosis than in thecontrol group of mices,and the pharmacological inhibitors of activation of NF-κB wasgiven can prevent I/R injury, reduce necrotic tumor necrosis factor-α (TNFα) andinterleukin-6(IL-6) releasing, inhibit inflammation and apoptosis, reduce infarct sizeand improve heart function. Myocardial ischemia and reperfusion are caused bymultiple factors. Therefore, the inhibition of JNK and its downstream NF-B may bethe treatment of myocardial ischemia-reperfusion injury.Berberine mainly is berberine medicine for the treatment of diarrhea, dysentery and other gastrointestinal infections in recent years. we have studied animalexperiments and clinical trials, and other findings proved that berberine cansignificantly improve cardiac systolic and diastolic function, stable cardiac electricalactivity and participation in myocardial energy metabolism. With the following studyof berberine found that berberine can protect myocardial I/R injury, reduce infarct size,and reduce the vascular endothelial injury, other studies show that berberine caninhibit inflammatory cytokine production, but the specific mechanism is unclear.Although berberine has much pharmacological activity, but its clinical application islimited by its poor absorption.Our made Huang Gui Solid Dispersion,Sodium caprateenhances intestinal absorption and increases intestinal permeability.Solid dispersioncan improve the solubility of poorly soluble drugs, significantly improve thebioavailability of berberine.However, Huang Gui Solid Dispersion whether to protectmyocardial ischemia reperfusion injury and its mechanism of action remains to befurther studied. Because berberine as main medicinal ingredient of HGSD, wespeculate that Huang Gui Solid Dispersion may be mediated by JNK and NF-κBsignaling pathway during ischemia/reperfusion, thereby inhibiting TNFα and IL-6release,reducing inflammation, might protect myocardial ischemia-reperfusion injury.Therefore, the present experiment will study its mechanism from two aspects,thewhole level and cell level.It is a theoretical providing and is basis for the treatment ofI/R injury in clinical applications.First, we have discussed the Huang Gui Solid Dispersion is able to againstmyocardial ischemia-reperfusion injury in the overall level.In the whole experiment, rats were randomly assigned to diffirrent groupstreatment group is administered HGSD,intragastric administration for7consecutivedays, doses of (12.5,25,50, IG) mg/kg.We establish isolated heart model ofischemia-reperfusion injury in rats, Control: continuous perfusion with KH solution105min. M group and HGSD administration model group: KH perfusion liquid stableafter30min, ischemia45min, reperfusion30min,and we detect data, including leftventricular developed pressure (LVDP), left ventricular end-diastolic pressure(LVEDP), left ventricular output (OUTPUT), the maximum rate of left ventricularpressure rise (+dp/dt) and the maximum rate of left ventricular pressure decreased(-dp/dt) changes. Enzyme troponin (cTnI) and cardiac tissue tumor necrosis factor(TNF-α), interleukin-6(IL-6) was detected..We assess tissue expression of JNK, NF-κB. Experimental results show that the model group ischemia45min, reperfusion30min, compared with the control group, M group LVDP, OUTPUT is reduced andLVEDP is elevated.cTnI is increased significantly, especially5min duringreperfusion.TNFα, IL-6has a significant increase. The expression of JNK, NF-κB isincreased.The LVDP, OUTPUT of HGSD administration group increases, LVEDP islowered,+dp/dt,-dp/dt increased in a dose-dependent manner, HGSD25, HGSD50administration was no different, so HGSD25as a follow-up dose experiments.Secondly in the cell experiments, we study Huang Gui Solid Dispersion onprotection mechanism of myocardial ischemia-reperfusion injury aboutinflammation-mediated JNK.Another intervent AMPK at different time points ofischemia or reperfusion to investigate the efficacy of HGSD main ingredient berberineactive role in the regulation of AMPK and JNK inhibition correlation betweeninflammatory pathway.First we prepare drug-containing serum,to detect berberine is used by HPLC.And calculate the mean serum berberine is27.95μM, with10%and20%serum forfurther study. Further we use the hypoxia chambers to copy hypoxia reoxygenationmodel, hypoxia4h,6h,8h, reoxygenation12h,and found that hypoxia8h, hypoxiareoxygenation12h,20%serum can reverse the injury, so subsequent experiments20%serum is of the therapeutic dose. In order to further study, JNK agonist(Anisomycin), inhibitors (SP6000125) is used alone or in combination with HGSDserum to observe Huang Gui Solid Dispersion serum how to protect H9c2cardiomyocytes hypoxia inducing inflammation injury in rats. It was found that theserum and JNK agonist group compared to serum group can significantly improvethe level of tumor necrosis factor (TNFα), interleukin-6(IL-6), but there was nosignificant difference between serum and JNK inhibitor (SP6000125)group; theserum and JNK agonist group also increased inflammatory injury of cell-associatedprotein expression and phosphorylation of JNK NF-κB nuclear translocation. Theresults suggest that HGSD can against serum inflammatory injury which caused byhypoxia, reduce the inflammatory cytokine release by inhibiting JNK phosphorylation,blocking NF-κB nuclear translocation.These results also indicate that serum HuangGui Solid Dispersion may improve hypoxia-induced inflammatory damage in H9c2,and reduce the inflammatory cytokine release by activation of JNK, inhibitingNF-κB phosphorylation. Moreover, the preliminary research has found that regulation of berberine onAMPK, the present study further investigated the active ingredient berberine ofHGSD to regulate AMPK and inhibit JNK pathway of inflammation.Through theestablish isolated rat heart model of ischemia-reperfusion injury, administration ofberberine (Ber) joint AMPK agonists during reperfusion period or administration ofberberine (Ber) joint AMPK inhibitors in ischemia to observe the datas. As shown, theleft ventricular developed pressure (LVDP) recovery rate,output OUTPUT (LVDP*HR)of Ber administration group,after45minutes of myocardial ischemia and30minutes reperfusion, significantly is improved.Left ventricular end-diastolic pressure(LVEDP) significantly is reduced. And reperfusion period is given AMPK agonists,ischemia is given AMPK inhibitors can counteract Ber group improved cardiacfunction. By western blot assay AMPK, JNK expression found berberine ischemiaand reperfusion in its different in AMPK regulation, and this ischemia and reperfusioninjury in the regulation of AMPK can inhibit JNK, indicating the active ingredientberberine of Huang Gui Solid Dispersion,regulation AMPK may be related to theinhibition of JNK pathways inflammation.In summary,Huang Gui Solid Dispersion can protect myocardialischemia-reperfusion injury in rats, JNK may be one of the key targets of itsprotective role, and its mechanism may be mediated by JNK NF-κB signalingpathway during ischemia/reperfusion, fillowing inhibit the release of TNFα andIL-6, reduce the inflammatory response.But the inhibition of JNK may be related tothe regulation of AMPK of berberine. |
