从兴奋收缩耦联机制看心力衰竭正性肌力药物发展

1会计学从兴奋收缩耦联机制看心力衰竭正性肌从兴奋收缩耦联机制看心力衰竭正性肌力药物发展力药物发展Sandow A (1952). Excitation-contraction coupling in muscular response. Yale J Biol Med 25 (3): 176201. PMID 130159500Bers, D. M. ExcitationContraction Coupling and Cardiac Contractile Force edn 2 (Kluwer Academic,Dordrecht, Netherlands, 2001).(Guinea-pig ventricular cell)Cardiac tissueT tubeCa+Ca+Ca+Ca2+PlbCa2+Ca+Ca2+Ca2+Ca2+Ca2+Ca2+Ca2+Ca2+Ca+Ca+Ca+Ca+Ca2+Ca+Ca+Ca+Ca+Ca2+Ca+Ca+Ca2+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca2+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca+Ca2+Ca2+Ca2+Ca2+Ca2+Ca2+Na+Na+Na+Ca2+SERCASRRyRL-Type Ca2+ChannelNa+/Ca2+ ExchangerCa+SarcolemmaCa2+ActinTropomyosinTroponinTitinMyosin Myosin-binding-protein C CapZ Tropomodulin Cross-linking protein 肌联蛋白（Titin）将粗肌丝与Z-线连接，维持肌原纤维的完整性和稳定性，保持舒张肌肉的静息张力，使粗肌丝处于肌小节的中央位置，使受牵拉的肌肉可恢复初始状态，以保证肌肉收缩时张力的输出。ZZTitin28,000 amino acids (3MDa) the largest protein known in mammals.TitinThin filament (Actin ,Tropom-yosin, Troponin) Thick filament (Myosin)Other proteinsChien, K.R., 1999F-actinZ-lineZ-lineThin Filament ProteinsG to F actin MW 42 kDaThe blue and grey molecules are actin monomers (MW 42.000)Ken C. Holmes: Max-Planck-Institute G-Actin F-Actin 肌动蛋白以两种形式存在, 即单体和多聚体。单体的肌动蛋白是由一条多肽链构成的球形分子, 又称球状肌动蛋白(globular actin, G-actin),外形类似花生果。肌动蛋白的多聚体形成肌动蛋白丝, 称为纤维状肌动蛋白(fibros actin, F-actin)。在电子显微镜下, F-肌动蛋白呈双股螺旋状, 直径为8nm, 螺旋间的距离为37nm。 Lorenz model of F-actin. A single G-actin monomer with inter-actin contact surfaces is shown on the right, the entire F-actin on the leftActin filaments are dynamic polymers whose ATP-driven assembly in the cell cytoplasm drives shape changes, cell locomotion and chemotactic migration. Actin filaments also participate in muscle contraction. The structure of the filament is not known at atomic resolution, but several models were produced in the laboratory of Ken Holmes (MPI for medical research, Heidelberg, Germany) by refinement against X-ray fiber diffraction dataTroponinHead-to-tail overlapABTakeda, S. et al. Nature 424, 35 41, 2003 HCTnCHCTnIHCTnTTropomyosinTropomyosinbinding regionHypervariable regionCrystal structure of human cardiac troponinC-DomainN-DomainCentral HelixEach TnC domain contains two motifs called EF hands, and it is the EF hands that directly bind calcium ions. Thus, the EF hands are TnCs way of sensing the calcium concentration; at 100 nM calcium (the usual cellular concentration) the N-domain EF hands are empty, but if the local concentration rises to 1 m mM, as it does when the muscle contracts, all of the EF hand bind calcium.KCa=3 x 105 M-1Ca2+-specificKCa= 2 x 107 M-1Ca2+-Mg2+ sites MYOSINMYOSIN MW 480 kDaForms thick filamentsHydrolyses ATPInteracts with F-actin 300-400 myosin molecules per 1 filamentS1S1150 nmMyosin重链重链 -helical coiled-coil轻链轻链160 nmS1S1 - Molecular Motor of Muscle ContractionRLCELCMyosin Head (S1) molecular motor of muscle contractionRLCELCATP Binding SiteActin Binding SiteATP (Myosin) ADP + Pi + EnergyF-actinCross-bridge Actin InteractionGordon et al. 2001A B C D EFrom Craig and Lehman, 2001,JMB 311, 1027The reversible binding of calcium to troponin alters the conformation of the thin filament, thereby turning muscle contraction ON and OFF Cross-bridge STATE: Thin filament STATE:Relaxed (OFF) BLOCKEDCa2+ Activated (Weak Binding) CLOSED Ca2+ and Myosin Activated (Strong binding) OPEN Three positions of Tropomyosin Activated Filaments (blue: actin bound end of actively cycling cross-bridges)Regulation of Muscle Contraction:/b/bATPCa2+Muscle ContractionPiIn the absence of Ca2+, the interaction of myosin with actin and consequently contraction is inhibited. Upon release of Ca2+ from the SR, the regulatory, Ca2+ specific sites of TnC bind Ca2+ exposing a patch of hydrophobic residues located in the N-terminal domain of TnC and the interaction of the TnC with TnI and TnT can take place. These internal Tn interactions promote translocation of the TnTm complex away from the outer domain of the actin filaments enabling the cyclic interaction between myosin heads (S1) and actin. The myosin head, an actin activated-Mg2+-ATPase dependent molecular motor, binds to actin and undergoes a power stroke, a phenomenon responsible for the interaction between the thick filament and the thin filaments and force generation. ATPase Cycle1. A M + ATP2. A+ MATP3. A M ADP Pi4. A M ADP +Pi5. A M +ADPPiADPPi release rate:10-20s-1Muscle Contraction Pi release rates:1. No Tm-Tn: 10 20 s-1 ; 2. + Tm-Tn no Ca2+: 0.1- 0.2 s-1 ; 3.+ Tm-Tn + Ca2+:10 20 s-1In vitro motility assay showing the sliding of actin filaments over a myosin surface initiated by flash photolysis of caged ATP(Clive R. Bagshaw)Bers DM. Cardiac excitation-contraction coupling J. Nature, 2002, 415(6868): 198-205.Excitation-contraction couplingRyanodine receptor(RyR) Phosphorylation of RYR increaseCa2+ leak ATP-dependent pump Phospholamban(PLB) In HF Expression and activation of SERCA2Phosphorylation of PLB Expression of 1ARATP supplyuptakeRe-uptake StoreRelease MSRSR Ca2+ srore decrease, Ca2+ transient delay123451. Reduced Ca+ trigger thru L-type channel2. Reduced RyR function (Calcium leaks from SR)3. Decreased sensitivity of TN-C to Ca+4. Reduced Ca+ uptake due to loss of SERCA function and increased Plb5. Increased Na/Ca exchanger functionDigilis purpureaDigilis purpureaPurple foxglovePurple foxgloveWilliam WitheringWilliam Withering (1741 - 1799)(1741 - 1799)DigitalisThe Effect of Digoxin on Mortality and Morbidity in Patients with Heart FailureN Eng1 Med,1997;336:525-533PlaceboDigoxinThe Effect of Digoxin on Mortality and Morbidity in Patients with Heart Failure N Eng1 Med,1997;336:525-53328%P0.01PlaceboDigoxinThe Effect of Digoxin on Mortality and Morbidity in Patients with Heart Failure N Eng1 Med,1997;336:525-533Opie, H. L. Drugs for the Heart. Orlando Florida: Grune & Stratton, Inc. 1980.DopamineDobutamineDies F, et al. Circulation 1986;74(suppl II):II-39. Uzunov, P. and Weiss, B Biochim. Biophys. Acta 284:220-226, 1972 Weiss, B. and Hait, W.N.: Ann. Rev. Pharmacol. Toxicol. 17:441-477, 1977. (according to Lippincott s Pharmacology, 2006)Packer M, et al. Effect of milrinone on mortality in severe chronic heart failure. N Engl J Med. 1991;325:1468-1475. ActinTropomyosinTnITnTCa2+cTnCMyosin head (S1 fragment)ATP pocketRLCELC左西孟旦左西孟旦Packer M. AHA Scientific Sessions, Dallas, USA, November, 2005.Packer M. AHA Scientific Sessions, Dallas, USA, November, 2005.33%30%White J ,Lee JA , Shah N , et al. Differential effects of the optical isomers of EMD53998 on contraction and cytoplasmic Ca2 + in isolated ferret cardiac mus-cleJ . Circ Res ,1993 ,73 :61270.Lee JA ,Allen DG. EMD53998 sensitizes the contractile proteins to calcium in intact ferret ventricular muscleJ . Circ Res ,1991 ,69 :9272936.Ribeiro da Silva MD, et al., Org Biomol Chem. 2004,2(17):2507-12.Ingraham RH, et al. Biochemistry,1988 Aug 9;27(16):5891-8.Tocchetti CG, et al. Circulation Res. 2007,100(1):96-104.Head-to-tail overlapPooled data of force development and Ca2+i transients at varied Ca2+o in the absence (open symbols) and presence (closed symbols)of AS (250m mmol/L).(P0.001 by one-way ANOVA).N=7 in each group.HNO不影响心肌的舒张功能不影响心肌的舒张功能The effects are HNO specific. The control drug (1-nitrosocyclohexyl tertbutylacetate) has no obvious effect on force of Skinning myocyte.Bers, D. M. ExcitationContraction Coupling and Cardiac Contractile Force edn 2 (Kluwer Academic,Dordrecht, Netherlands, 2001).Myosin Head (S1) molecular motor of muscle contractionRLCELCATP Binding SiteActin Binding SiteATP (Myosin) ADP + Pi + EnergyMuscle Contraction Pi release rates:1. No Tm-Tn: 10 20 s-1 ; 2. + Tm-Tn no Ca2+: 0.1- 0.2 s-1 ; 3.+ Tm-Tn + Ca2+:10 20 s-1Re-uptake StoreRelease MSRSR Ca2+ srore decrease, Ca2+ transient delayWhite J ,Lee JA , Shah N , et al. Differential effects of the optical isomers of EMD53998 on contraction and cytoplasmic Ca2 + in isolated ferret cardiac mus-cleJ . Circ Res ,1993 ,73 :61270.Lee JA ,Allen DG. EMD53998 sensitizes the contractile proteins to calcium in intact ferret ventricular muscleJ . Circ Res ,1991 ,69 :9272936.Head-to-tail overlap