Intervention Of Scutellaria Baicalensis Stem-leaf Total Flavonoids On Myocardial Ischemia And Reperfusion JAK/STAT Pathway In Rats

Myocardial ischemia reperfusion injury is a familiar clinical pathologicalprocess, and is a main interference in the reperfusion therapy for acute myocardialinfarction. The mechanism is still not very clear completely. In recent years，agreat deal of studies have been carried out on this kind of injury and the resultsshow apoptosis contributes to myocardial cell death during ischemia reperfusioninjury.Some experiments have indicated that JAK (just anotherkinase)/STAT(signal transductors and activator of transcription）pathway play avery important role on the myocardial apoptosis induced by ischemia reperfusioninjury.Objective:The study used the model of myocardial ischemia reperfusion injury byligating the left descend coronary artery to mimic the pathological process， inwhich the human cardial tissue damage was made by exsanguine-reperfusion fromthe thrombolysis therapy，to observe，on the whole level，the protective effectsof Scutellaria Baiculensis Stem-leaf Total Flavonoid on myocardial cell during theischemia reperfusion process in rats，and whether the mechanism is achievedthrough activation of JAK/STAT pathway,to provide accurate experimental andtheoretical evidence in order to exploit the new drugs to prevent and treat theischemic heart disease.Methods:50adult SD(Sprague dawley) rats(weight220±20g) were divided at randominto SSTF pretreatment Ⅰg roup、SSTF pretreatment II group、SSTF pretreatmentIII group、ischemia reperfusion group and sham operation group，there are ten ratsin every group.Rats in SSTF pretreatment group were perfused withSSTF(17.5mg/kg/d，35mg/kg/d，70mg/kg/d respectively) a week before operation， and to the sham operation group and the ischemia reperfusiongroup，saline was given instead. Myocardial ischemia reperfusion injury modelwas made by ligating the left descend coronary artery30minutes andreperfusing2hours. ie, Besides Sham group, other groups were subjected to30minutes of left anterior descending coronary artery (LAD) occlusion and followedfor2hours of reperfusion. Apoptosis of myocardial cells were detected by flowcytometry.Cardiocyte ultrastructural pathological changes were observed withtransmission electron microscope. The immunohistochemical method was usedto determine the expression of JAK2,STAT1and STAT3gene.Results:1Apoptosis of myocardial cells were detected by flow cytometryCompared with that in sham operation group(6.78±0.96)%，apoptosis rate ofmyocardial cell is higher significantly in ischemia reperfusion group（12.45±2.17）%,(P＜0.01）;however, apoptosis rate of myocardial cell is lowerobviously in SSTF pretreatment group（10.32±2.04，9.44±1.85，7.96±1.23）%than that in ischemia reperfusion group (P＜0.05，P＜0.01）. SSTFⅡ and SSTFⅢ compared to SSTFⅠ, apoptosis rate of myocardial cell have significantdifference(P＜0.01), but the results of two group(between SSTFⅡ and Ⅲ group)have no significant difference (p>0.05). SSTF pretreatment groups showed a dosedependent effect.2Ultrastructure alteration of myocardial cells①Sham group：myocardial ultrastructural changes slightly in ischemiccenter、marginal zone and non-ischemic zone. The structure of muscle filaments isvisible;sarcomere is consistent;the light and dark belt of muscle fiber isclear;sarcolemma integrity;mitochondrial membrane integrity and ridge gapwidened slightly.②IR group：myocardial cell swelled obviously,sarcolemmadamaged,chromatin margined;muscle fiber reduced largely;a large number offilaments broken and dissolved and form ed muscle lesions varyingsizes;mitochondria swelled,crestis irregularor broken into flocculent, appearedempty bubble-based; sarcoplasmic reticulum expanded;myocardial cells apoptosisbody appeared③SSTFⅠgroup: myocardial cell swelled in ischemic center; local sarcolemma fractured; part of muscle fiber arranged irregularly; muscle lesionswas less than that in IR group; mitochondria swelled,but the outer membranealmost integritied，ridge paralleled or broken into flocculent; empty bubble-basedmitochondria existed too.④SSTFⅡ group：broken and dissolved muscle filamentsstill existed; muscle fiber in ischemic center、marginal zone arranged regularly;mitochondria swelled slightly， outer membrane integritied，electron density inmost of the mitochondrial matrix increased, some mitochondrial ridge rupturedmildly.⑤SSTFⅢ group：muscle fiber in different zone arranged neatly，A、Z、H belt cleared，mitochondria swell reduced。3the expression of JAK2protein of myocardium by immunohistochemis-tryThe JAK2protein in sham operation group，ischemia reperfusion group，SSTF pretreatment Ⅰ,II and III group were respectively（0.096±0.018）%、（0.278±0.049）%、（0.254±0.047）%、（0.223±0.036）%、（0.198±0.025）%；which indicated that JAK2protein expression was significantly more in ischemiareperfusion group than that in sham operation group(P＜0.01)，but significantlyfewer in SSTF group than that in ischemia reperfusion group（P＜0.05，P＜0.01）.SSTFⅡ and SSTFⅢ compared to SSTFⅠ, the expression of JAK2protein wasmarkly lower than SSTFⅠ, have significant difference(P＜0.01).4the expression of STAT1and STAT3protein of myocardium byimmunohistochemistryThe STAT1protein in sham operation group，ischemia reperfusion group，SSTF pretreatmentⅠ、II and III group were respectively （0.084±0.05）%、（0.257±0.09）%、（0.211±0.07）%、（0.168±0.04）%、（0.117±0.06）%；which indicated that STAT1protein expression was significantly more in ischemiareperfusion group than that in sham operation group(P＜0.01)，but significantlyfewer in SSTF group than that in ischemia reperfusion group（P＜0.05，P＜0.01）.Compared with SSTFⅠgroup, STAT1protein expression was significantlydecreased in SSTF II and III group(P＜0.01).The STAT3protein expression in sham operation group，ischemia reperfusiongroup，SSTF pretreatmentⅠ，II and III group were respectively （0.276±0.09）%、（0.074±0.04）%、（0.185±0.06）%、（0.193±0.08）%、（0.237±0.10）%,which indicated that STAT3protein expression was significantly fewer in ischemiareperfusion group than that in sham operation group(P＜0.01)，but significantlymore in SSTF groups than that in ischemia reperfusion group （P＜0.05，P＜0.01）. Compared with SSTFⅠgroup, STAT3protein expression was significantlyincreased in SSTF II and III group(P＜0.01). But between SSTF II and SSTF IIIgroup have no significant difference (p>0.05).Conclusions:1. Prophylactic use of SSTF can improve the ability of myocardium onresisting ischemia reperfusion injury, and reduce the damage degree of myocardialultrastructures.2Prophylactic use of SSTF can decrease apoptotis of cardiomytocytes, andthe effect in the SSTFⅡ、Ⅲ group is better than SSTFⅠgroup. SSTF have aprotective effect on the myocardial tissue injury caused by ischemia reperfusionthrough up–regulating protein expression of STAT3and down-regulating proteinexpression of JAK2, STAT1.To sum up, SSTF play the protective role to the myocardium throughactivating the JAK/STAT pathways.