【作者】 罗义；

【导师】 刘伊丽； 陈瑗；

【作者基本信息】 第一军医大学 ， 心血管内科学， 2000， 博士

【摘要】 研究目的 顿抑心肌中氧自由基生成增加，氧自由基抑制心肌收缩功能，这两点是明确的。但氧自由基是靠直接的细胞毒作用还是通过形成其它产物来引起心肌顿抑，目前仍未明了。最近研究提示，一氧化氮（NO）与超氧阴离子（·O₂^-）反应生成的过氧亚硝酸阴离子（ONOO^-）在·O₂^-启动的细胞损害中起着重要作用。此外，顿抑心肌是否存在微循环障碍，这是一个尚未解决的问题。本研究的目的是：1.探讨顿抑心肌微循环改变及机制；2.探讨NO和ONOO^-在心肌顿抑发生中的作用及机制；3.观察NO合成酶抑制剂和云芝多糖（PSK）对顿抑心肌结构、功能及微循环的影响。 材料和方法 39只雄性杂种犬随机分入6组。1.短时间缺血顿抑（MSS）组：左前降支冠脉（LAD）结扎15min、再灌注120min；2.长时间缺血顿抑（MSL）组：LAD结扎60min、再灌注120min；3.N^ω-硝基-左旋精氨酸甲酯（NAME）组：MSL加NAME干预；4.氨基胍（AMD）组：MSL加AMD干预；5.云芝多糖（PSK）组：MSL加PSK干预；6.假手术（SHAO）组。全过程监测血流动力学和心电图。在不同观察时间点静脉注射含全氟丙烷声振白蛋白微泡造影剂，采用二次谐波成像和间歇发射技术行心肌声学造影（MCE）。由主动脉根部分别注射乙酰胆碱（ACH）和硝酸甘油（NG）后即刻重复MCE以检测冠脉舒张功能。每次MCE后行超声心功能测定。相应时间点冠状静脉窦抽血测定血浆乳酸、NO和丙二醛浓度。心肌标本行病理检查。 以MCE心肌视频密度峰值（PVI）表示心肌血流灌注，顿抑区与正常区PVI比值（PVIR）表示顿抑区相对血流灌注。MCE曲线上升斜率和曲线早期下降斜率分别代表心肌血流灌注速度和排空速度。以注射ACH或NG后、前顿抑心肌PVI比值分别表示心肌微血管内皮依赖性舒张（EDR）功能和非EDR功能，以注射ACH或NG后、前顿抑心肌PVIR比值分别表示相对性EDR功能和相对性非EDR功能。 结果1．血流动力学改变：除NAME组再灌注期心率减慢、平均主动脉压升高外，其余各组心率和平均主动脉压无显著变化；MSS组、MSL组和NAME组再灌注前和再灌注早期左室舒张压显著升高，AMD组和PSK组仅再灌注前左室舒张压升高。2．心肌血流和微血管舒张功能改变：MSS组和MSL组再灌注早期心肌充血，血流灌注速度和排空速度加快；微血管EDR和非EDR功能减弱，其恢复速度与原缺血时间长短有关：＊＊＊E显著减慢再灌注期心肌血流灌注速度和排空速度，引起心肌缺血，加重微血管EDR和非EDR功能损害：AMD和PSK减轻再灌注期心肌充血程度，使再灌注期加快的血流灌注速度和排空速度．减慢，促进微血管EDR和非EDR功能的恢复。3．心功能改变：MSS组和MSL组再灌注期心肌节段收缩期增厚百分率和左室射血分数显著下降，随着再灌注时间的延长心功能呈进行性改善；NAME加重心功能损害；AMD和 PSK促进再灌注期心肌功能的恢复。4．血生化指标变化：＊)乳酸：MSS组再灌注早期乳酸浓度显著升高，至再灌注120min恢复正常；MSL组和NAME组再灌注前乳酸浓度即明显升高，再灌注早期进一步升高，至再灌注120min尚未恢复正常；AMD组和PSK组再灌注前和再灌注早期乳酸浓度升高，至再灌注 120min恢复正常。口)NO：MSS组和 MSL组再灌注期NO浓度明显升高，至再灌注 120min 尚未恢复正常：＊＊＊E组整个再灌注期＊O浓度明显减低；＊＊D组和PSK组再灌注期NO浓度轻度升高，但与结扎前相比，除PSK组再灌注30min时差异有统计学意义外，其余均无统计学意义。（3）丙H醛：MSS组、MSL组和 NAME组再灌注期丙二醛浓度明显升高，至再灌注 120min 尚未恢复至结扎前水平。AMD组和PSK组再灌注早期丙二醛浓度升高，但恢复较快。5．顿抑心肌病理改变：MSS组心肌细胞除偶见线粒体轻度脱颗粒外，未见其它超微结构改变。MSL组心肌组织水肿，毛细血管内皮细胞稍肿胀，内皮细胞间连接间隙稍增宽；心肌细胞部分肌丝断．裂，收缩带溶解，线粒体肿胀、脱颗粒，胞质水肿。NAME组心肌组 织水肿，毛细血管内皮细胞显著肿胀，内皮细胞间连接间隙明显增宽， 基底膜电于密度减低；心肌细胞肌丝断裂，收缩带溶解、变模糊或消失， 线粒体明显肿胀、脱颗粒，线粒体脊断裂，胞质水肿：心肌细胞间连接 间隙增宽，闰盘结构变模糊。AMD组和PSK组心肌组织轻微水肿，毛 ．－仁 细血管和内皮细胞结构未见明显改变，心肌细胞除偶见胞质轻微水肿、 线粒体肿胀和脱颗粒外，未见其它明显异常改变。硝基酪氨酸免疫组化 检查发现，MSS组顿抑心肌组织见较强阳性染色的心肌细胞灶；MSL 组顿抑心肌组织见心肌细胞强阳性染色，主要是胞浆尤其横纹处染色较 深，阳性染色的心肌细胞呈灶性分布，而且阳性灶较大、较多：＊＊＊E 组顿抑心肌组织中偶见单个心肌细胞局部弱阳性染色；AMD组和pSK 组顿抑心肌组织偶见心肌细胞较弱阳性染?更多还原

【Abstract】 Mechanisms of the pathogenesis of myocardial stunning are not fully understood. It has been known that reactive oxygen species(ROS) production increases in myocardial stunning and ROS depress myocardial function. But it is unclear whether ROS impair myocardium directly or through the formation of other substances. Recently, it has been demonstrated that ischemialreperfusion(I/R) leads to the activation of nitric oxide synthase (NOS) and nitric oxide(NO) participates in the regulation of hemodynamic and microcirculatory changes in organic hR. The dual role of NO as a cytoprotective or a cytotoxic free radical gas has been noted in various types of pathophysiological conditions. However, It is controversial whether NO is protective or deleterious against myocardial hR injury? Recent studies suggest that peroxynitrite(ONOO ) formed by the reaction of NO with superoxide plays an important role in the cell damages caused by superoxide. Additionally, there is a continuing debate as to the effects of inhibitors of NOS on myocardial hR injury. Finally, whether intramyocardial microcirculatory dysfunction occurs in stunned myoeardiurn is an unresolved issue. The purposes of this study were (1) to research the intramyocardial microcirculatory derangement and its mechanism during myocardial stunning, (2) to study the roles of NO and ONOO in the pathogenesis of myocardial stunning and their mechanisms, (3) to evaluate the effects of a nonselective NOS inhibitor, N -nitro-L-arginine methyl ester(NAME), a selective inducible NOS(iNOS) inhibitor, aminoguanidine (AMD), and an antioxidant, polysaccharide krestin(PSK), on microcireulatory changes, myocardial performance and structural damages after myocardial stunning. Methods Thirty-nine adult mongrel dogs of male, weighing 13-18 kg, anesthetized with sodium pentobarbital, were randomly assigned to six groups. (I )Myocardial stunning after a short-time ischemia(MSS) group: dogs underwent 15 mm of left anterior descending coronary artery(LAD) occlusion, followed by 120 mm reperfusion. (2)Myocardial stunning after a long-time ischemia(MSL) group: dogs underwent 60 mm of LAD occlusion, followed by 120 mm reperfusion. (3)NAME group: dogs underwent the same hR as MSL group, received NAME(lOmg/kg). One-third of NAME dose was given intravenously 10 mm prior to LAD occlusion, and the rest was infused continuously intravenously from 10 mm before reperfusion to the end of reperfusion. (4)AMD group: dogs were subjected to the same I/R as MSL group, received AMD(l0Omg/kg). AMD was given by the same way as NAME. (5)PSK group: animals were subjected to the same J/R, received PSK. which was administered orally, at a dose of 150mg/kg once daily for two day抯 before thoracotomy. (6)Sham operation(SHAO) group. Myocardial contrast echocardiography (MCE) was performed using intravenous perfluoropropane-exposed sonicated dextrose albumin(0.Olml/kg) at baseline, during coronary occlusion, and at 5,30,60.90.and 120 mm after reperfusion. At baseline. 5.60,and 120 mm after reperfusion. acetylcholine (ACH, 10 1-? g/kg) and nitroglycerin(NG. 10 p g/kg) were given from aortic root injections after resting MCE, respectively, and MCE was repeated immediately, respectively. Electrocardiogram-gated end-systolic images in short axis were acquired in harmonic mode and digitized on-line. Left ventricular function was evaluated by?echocardiography using an echocardiogram at the above time points. Hemodynamics was measured by更多还原