心肌缺血预处理ppt课件

Click to edit Master title style,Click to edit Master text styles,Second level,Third level,Fourth level,Fifth level,*,EARLY SIGNALING PATHWAY IN HEART PROTECTION BY ISCHEMIC PRECONDITIONING,EARLY SIGNALING PATHWAY IN HEA,1,Ischemic preconditioning (PC) is the,phenomenon describing protection of,the heart against reperfusion injury,following prolonged ischemia,by virtue of,prior,short episode,(or episodes) of ischemia and,reperfusion.,Ischemic preconditioning (PC),2,An analogous phenomenon of cardiac protection against reperfusion injury by pre-administration of a drug is called,drug-induced PC,.,We have developed a family of drugs that can do it.,An analogous phenomenon of car,3,Ischemia,R,1,R,2,Stabilization,10 min,Reperfusion,20 min,Pc,1,Pc,2,Pc,3,R,3,The System:,Isolated rat heart,The procedure :,Three episodes of 2 globalischemia separated by 3 reperfusion,PC,IschemiaR1R2StabilizationReper,4,Two windows of protection by ischemic PC have been identified: 1 3 hours following PC, and 24 hour following PC.,We work with the isolated rat heart, and will focus on the first window.,Two windows of protection by i,5,Cardiac,Function,without and with PC has been studied. Below the following parameters are presented:,DP = PSP - EDP,+dP/dt,EDP,other parameters not shown,Cardiac Function without and w,6,Developed Pressure (DP) (% of pre-ischemic value),Versus,Ischemic Time Preconditioning (PC),Developed Pressure (DP) (% of,7,+dP/dt (%),Versus,Ischemic Time Preconditioning (CPC),+dP/dt (%) Versus Ischemic Tim,8,EDP (mmHg),Versus,Ischemic Time Preconditioning (CPC),EDP (mmHg) Versus Ischemic Tim,9,BIOCHEMISTRY,BIOCHEMISTRY,Biochemical parameters of the heart tissue: a similar protection was observed for ATP, energy charge, ascorbate and other parameters.,in the coronary flow: leakage into the CF of cellular metabolites:,BIOCHEMISTRYBIOCHEMISTRY,10,HPLC Analysis of the First 10-CFFsFollowing 35 Global Ischemia,PC,0,10,20,30,40,50,60,70,Control: ischemia of 35- no PC,Preconditioning followed by 35 ischemia,S,A,S,P,UA,Ino,AA,HPLC Analysis of the First 10-,11,Currently Proposed Mechanisms of Protection by PC,The detailed understanding of the mechanism of PC is still under investigation.,Several possibilities have been suggested including:,adenosine activation;,preservation of high-energy phosphates;,washout of metabolites;,role of Ca,+,-channels;,Currently Proposed Mechanisms,12,increase in antioxidants activity;,synthesis of heat-shock proteins, G-proteins;,nitric oxide production;,role of K,+,-channels;,role of PKC,WE HAVE BEEN STUDYING THE ROLES OF IRON, COPPER AND FREE RADICALS IN ISCHEMIA AND OBTAINED INDICATIONS FOR THEIR INVOLVEMENT IN CARDIAC PC,increase in antioxidants activ,13,ROS - Reactive Oxygen Species - in reperfusion injury,The,formation of ROS,from relatively less reactive species is,mediated by redox-active,metal ions.,Circumstantial evidence suggests a,causative role,for newly mobilized redox-active iron in tissue injury.,ROS - Reactive Oxygen Species,14,Two lines of evidence support this idea:,Inclusion of,iron chelators,in the perfusate,has been shown to protect against tissue injury following ischemia.,Addition of,iron or copper,to the perfusate,facilitated injury in hearts subjected to ischemia and reperfusion.,Two lines of evidence support,15,McCords Scheme for Myocardial Damage Following Ischemia and Reperfusion,reoxygenation,Ischemia,McCords Scheme for Myocardial,16,McCord scheme,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,McCord scheme Ischemia reox,17,McCord scheme,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,Xanthine,dehydrogenase,Xanthine,oxidase,McCord scheme Ischemia reox,18,McCord scheme,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,Xanthine,dehydrogenase,Xanthine,oxidase,Ca,2+,Protease,McCord scheme Ischemia reox,19,McCord scheme,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,Xanthine,dehydrogenase,Xanthine,oxidase,O,2,Ca,2+,Protease,O,2,McCord scheme Ischemia reox,20,McCord scheme-modified,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,Xanthine,dehydrogenase,Xanthine,oxidase,O,2,Ca,2+,Protease,O,2,OH,McCord scheme-modified Ischem,21,McCord scheme -modified,Ischemia,reoxygenation,ATP,AMP,Adenosine,Inosine,Hypoxanthine,Xanthine,dehydrogenase,Xanthine,oxidase,O,2,Ca,2+,Protease,O,2,OH,Cu/Fe,McCord scheme -modified Ische,22,Fe(II) + H,2,O,2,Fe(III) + OH,-,+ OH,Fenton Reaction,Fe(II) + H2O2 Fe(III) + OH,23,Haber Weiss Reaction,or,Superoxide-,Driven Fenton Reaction,O,2,+ Fe(III) O,2,+ Fe(II) (1),Fe(II) + H,2,O,2,Fe(III) + OH,-,+ OH(2),O,2,+ H,2,O,2,OH + OH,-,+ O,2,(3),-,-,Haber Weiss Reactionor Super,24,Fe and Cu,Fenton and Haber-Weiss reactions for copper are also valid.,copper might prove even more important than iron in catalyzing such a reaction, and as mediators of tissue injury (,conc, mobility and reactivity,).,Fe and Cu,25,Cardiac injury well correlated with the level of Fe/Cu mobilization from the heart tissue to the coronary flow (CF). It was further proposed that Fe/Cu levels in the CF can serve as predictive criteria for the degree of cardiac injury,心肌缺血预处理ppt课件,26,MOBILIZATION OF Fe/Cu into CORONARY FLOW FRACTIONS,CFF#1,versus,TOTAL Fe/Cu,0,20,40,60,80,100,120,140,18,35,60,CFF #,9,8,7,6,5,4,3,2,1,0,Fe (ng/ml),MOBILIZATION OF Fe/Cu into COR,27,TOTAL,Fe,levels in the CFFs #1 Versus Ischemic Time Preconditioning,70,60,50,40,30,20,10,0,20,40,60,80,100,120,140,Control,PC,Duration of Ischemia,Fe (ng/ml),TOTAL Fe levels in the CFFs #1,28,TOTAL Cu,levels in the CFFs #1 Versus Ischemic,Time Preconditioning,70,60,50,40,30,20,10,0,10,20,30,40,50,Control,PC,Duration of Ischemia,Cu (ng/ml),TOTAL Cu levels in the CFFs #1,29,心肌缺血预处理ppt课件,30,Isolated Rat Heart (Langendorff),R,1,R,2,Isch,Time:,10 2 3 2 3 2 3,18 - 60,20,Stabil,Reperfusion,Pc,1,Pc,2,Pc,3,R,3,CFF# 0 1, 2 1, 2 1, 2 1.10,Collection,time: -10 3 0 3 0 3 0 - 1,Isolated Rat Heart (Langendorf,31,Iron in the CFFs following Global Ischemia,without PC,0,20,40,60,80,100,120,140,18,35,60,CFF #,9,8,7,6,5,4,3,2,1,0,Fe (ng/ml),Iron in the CFFs following Glo,32,Iron in CFFs FollowingPC and Global Ischemia,0,35,70,105,140,CFF #,0 1 2 1 2 1 2,pc1 pc2 pc3,1 2 3 4 5 6 7 8 9 10,Iron in CFFs FollowingPC and,33,Copper in the CFFs following Global Ischemia,10,9,8,7,6,5,4,3,2,1,0,0,10,20,30,40,CFF#,18,35,60,Cu(ng/ml),Copper in the CFFs following,34,Copper in The CFFs Following Preconditioning and Global Ischemia,0,10,20,30,40,CFF #,pc1 pc2 pc3,1 2 3 4 5 6 7 8 9 10,0 1 2 1 2 1 2,Cu (ng/ml),Copper in The CFFs Following,35,Ischemia 18,Ischemia 35,Ischemia 60,0,1-1,1-2,2-1,2-2,3-1,3-2,40,20,10,0,30,0,40,80,120,160,1,2,3,4,5,6,7,8,9,10,1,2,3,4,5,6,7,8,9,10,0,40,80,120,160,0,1-1,1-2,2-1,2-2,3-1,3-2,0,40,80,120,160,1,2,3,4,5,6,7,8,9,10,40,20,10,0,30,1,2,3,4,5,6,7,8,9,10,40,20,10,0,30,Cu (ng/ml),Fe (ng/ml),Ischemia 18Ischemia 35Ischem,36,TOTAL Fe/Cu,versus,LAMP (LIP),Is it the,total,Fe/Cu that plays a role in,mediation of injury?,“LAMP”,- the,LA,bile transition,M,etals,P,ool is the active fraction, and,is important!,The over-estimated values in the literature,How can we quantitate LAMP?,心肌缺血预处理ppt课件,37,CFF-LAMP,Mediated Conversion of Salicylate to 2,5-DHBA,70,60,50,40,30,20,10,0,10,20,30,40,50,Control,PC,Duration of Ischemia,2,5-DHBA (ng/ml),CFF-LAMP Mediated Conversion o,38,CFF-LAMP,Mediated Conversion of Salicylate to 2,3-DHBA,70,60,50,40,30,20,10,-5,5,15,25,35,45,55,Control,PC,Duration of Ischemia,2,3-DHBA (ng/ml),CFF-LAMP Mediated Conversion o,39,Three levels of mobilized,metal ions:,PC phase,prolonged ischemia following PC,prolonged ischemia,Three levels of mobilized,40,Ferritin Levels in Heart Tissue and in Coronary Flow (CF) following 35 ischemia,HEART TISSUE,Total Ft in,Ft/ProteinFe/Protein,CF,(,m,g/mg) (,m,g/mg),(,m,g),Ischemia/reperfu-,sion group,(,80,),0.54,+,0.07 0.167 1.30,+,0.14,PC/Ischemia and,Reperfusion group,(80) 0.90,+,0.13* 0.198 0.73,+,0.11,Normal-Perfusion,group (80),0.64,+,0.10 0.196 0.19,+,0.05,Ferritin Levels in Heart Tissu,41,Iron Level in Heart Tissue Ferritin,(,m,g/,m,g),Experimental GroupFe in Ferritin,(,m,g/,m,g),Ischemia group (80 min)0.31,+,0.04,PC group (80 min)0.22,+,0.04*,Normal perfusion group (80),0.32,+,0.04,PC (25 min)0.30,+,0.01,Iron Level in Heart Tissue Fer,42,Three levels of mobilization of iron: high levels are associated with tissue injury.,The PC phase: low levels of mobilized iron (and copper) and no damage.,Removal of the inhibitory control of protein synthesis through the conversion of IRP to Aconitase.,Three levels of mobilization o,43,Ferritin,Ferritin is an intracellular iron binding protein, able to store about 4,000-4,500 iron atoms per molecule. Under normal conditions the amount of iron in heart Ferritin is about 1,500-2,500 iron atoms.,FerritinFerritin is an intrac,44,Ferritin,Sullivan (1981) and Salonen et al. (1992) reported an association between the increase of,iron stores,and the,risk of myocardial infarction,. They demonstrated an important role for the increase of ferritin in ischemic heart disease. Magnusson et al. (1994) and Enbergs et al (1998), on the other hand, did not find such a correlation between ferritin levels and myocardial infarction.,Recent publications (1997 - 2000) tend to support the,claim,that ferritin is a marked risk factor for AMI.,FerritinSullivan (1981) and Sa,45,Does Ferritin Synthesis take place,during or following PC ?,Is protein synthesis required for PC?,Does it involve transcriptional or translational control?,Does Ferritin Synthesis take p,46,Ferritin synthesis,White and Munro (1987) reported about,rapid translational response,to iron in rat liver: mRNA of both ferritin subunits was mobilized to polysomes.,Tacchini et al. (1997) and Cairo et al. (1998) showed that,ischemia-reperfusion,and,oxidative stress,lead to inhibition of,IRP,activity followed by activation of ferritin synthesis.,Ferritin synthesisWhite and Mu,47,Several studies have indicated that protein synthesis is required,for PC. The,PC effect was abolished only with cycloheximide,but not with actinomycin D, indicating that PC is regulated at the,post-transcriptional level.,In another study:,Inhibitors of protein synthesis have been used in the,in vivo,rabbit,model.,Neither cycloheximide nor actinomycin D,did,reverse the,PC effect. The inhibition of protein synthesis was not complete,(85%) and only one episode was applied (Thornton et al., 1990).,Several studies have indicated,48,Our preliminary results indicate that PC is associated with,accumulation,of ferritin, and fits into the,post-transcriptional,regulation model.,Translation of Ft could occur by taking advantage of Ft-mRNA,that is present in the cells.,Our preliminary results indic,49,Signaling,with “LAMP” in PC,During the PC phase,small but reproducible,levels of Fe/Cu are liberated from their proteins, and serve as intracellular and/or intercellular signals for cascade of events.,For,iron,- the signal could be transduced through IRP, removing the inhibition of translation of ferritin.,Signaling with “LAMP” in PCDur,50,The proposed mechanism of PC protection:,The small levels of Fe (and Cu?) mobilized during/following the PC serve as signaling species reverting the inhibitory control of ferritin synthesis in heart cells, and are not involved in tissue damage.,The high levels of Fe/Cu following prolonged ischemia serve as catalysts that are involved in the injurious process to the heart. PC yields more apo- ferritin that can scavenge newly mobilezed iron.,The proposed mechanism of PC p,51,Is,Copper,Important in PC?,Recently published data have indicated important roles for,metallothionein (MT),in reperfusion injury and PC.,Hearts from transgenic mice that overproduced MT were better protected against ischemia (as indicated by infarct size, hemodynamic function and leakage of proteins), than hearts from non- trangenic animals (Kang, Li & Saari, 1999),Is Copper Important in PC?Rece,52,Ischemic-PC,of the brain was found to provide protection against edema and,neuropathology, in kainic acid-induced status epilepticus. This protection was dependent on protein synthesis (Emerson et al., 2000). Analysis of MT-1, MT-2 and heme,oxygenase-1 (HO-1) revealed that the,PC had induced a pattern of induction,and suppression of these genes, that have,led to an adaptive response, preventing the,seizure-associated neuropathology.,Ischemic-PC of the brain was f,53,Conclusion:,We,distinguish,between the burst of Fe/Cu in the early reperfusion phase following prolonged ischemia and the small amounts of Fe/Cu liberated/mobilized during or immediately following PC.,PC,leads,to the mobilization of low levels of iron and copper, that in turn, serve as signals, inducing synthesis of their storage proteins, and leading to the scavenging of excess metals and the inhibition of transition metal-mediated tissue injury.,Conclusion:,54,