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MMP in the failing heart: a potential therapeutic target

MMP in the failing heart: a potential therapeutic target. Heart failure. Heart failure is a major public health problem (1% of the total population in France).

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MMP in the failing heart: a potential therapeutic target

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  1. MMP in the failing heart: a potential therapeutic target Heart failure Heart failure is a major public health problem (1% of the total population in France). It is is a common sequel to many forms of heart diseases, with various causes including pulmonary disease, myocardial infarction, arrhythmias, anemia, renal failure, nephrotic syndrome, and thyroid disease. It is a complex disease, with many phenotypes, but which displays two main characteristics: cardiac growth and cardiac remodeling.

  2. Cardiac remodeling in heart failure Normal heart. Hypertrophic heart. Failing heart. Large left ventricular chamber, thin left ventricular wall, loss of pumping capacity. Thick walls, small ventricular chambers, increased pumping capacity. (Freeman et al. JCI, 2001, 107:967; Limbird and Vaughan, PNAS, 1999, 96: 7125) The remodeling process is not only due to changes in the cardiac cells but also to changes in the extracellular matrix.

  3. Extracellular matrix (ECM) in the heart Scanning electron micrograph, longitudinal and transversal sections, showing collagen architecture in the left pig heart ventricle.(Zenat et al. Am J Physiol (2000) 279:H602) The myocardial ECM contains fibrillar collagens (types 1 and 3) which ensure structural integrity of the adjoining myocytes and alignment which imparts mechanical support to the myocardium and govern tissue stiffness. Disruption within the ECM network will lead to loss of a normal structural support and abnormal stress for cardiomyocytes during contraction. This will result in changes in myocardial geometry and function.

  4. Matrix metalloproteases (MMPs) in the heart MMP-2 , MMP-9 , MMP-3, MMP-1 , MMP-13 and MT1-MMP have been identified in the human heart, and are responsible for ECM degradation. A cause/effect relationship between MMPs and the left ventricular remodeling proces in the course of heart failure has been documented by several studies using different approaches: - expression and activity of the cardiac MMPs in animal models of heart failure, and human failing hearts; - studies in knocked-out and transgenic animal models; - pharmacological studies with MMP inhibitors.

  5. Rapid pacing leads to left ventricular dysfunction. Expression and activity of the cardiac MMPs increase time-dependently, together with the progression of heart failure. Immuno blotting Gelatin zymography Expression and activity of MMPs in heart failure (1). Rapid pacing in pig. (Spinale et al. Circ.Res., 1998,82:482) In this model, collagen content in the heart decreases.

  6. Non infarcted infarcted Expression and activity of MMPs in heart failure (2). The infarcted mouse. In the infarcted mouse, cardiac MMPs expression is increased. Ducharme et al. JCI, 2000, 106:55 Increased MMPs surpass TIMPs and this occurs together with an augmentation in the collagen content. Immunoblotting

  7. 6 weeks 11 weeks 15 weeks Iwanaga et al. JACC, 2002, 39:1384 DS: Dahl salt-sensitive rat DR: Dahl salt-resistant rat Gelatin zymography Immunoblotting Expression and activity of MMPs in heart failure (3). The Dahl salt-sensitive rat. The Dahl salt-sensitive rat develops hypertension associated with a rapidly developing heart failure when it is given a diet supplied with sodiumsalt. In the DS rats, MMP-2 activation surpasses that of TIMPs. Myocardial fibrosis appears focally.

  8. Expression and activity of MMPs in heart failure (4). The human failing heart. Li et al. Circulation 1998, 98:1728 Thomas et al. Circulation 1998, 97:1708 Immunoblottings Gelatin zymographies MMP-3 and MMP-9 are increased. TIMP-1, -3 and -4 are reduced. Such an imbalance favors persistent MMP activity.

  9. Heling et al. Circ Res 2000, 86:846-853 Expression and activity of MMPs in heart failure (5). The human failing heart. The increased content in MMPs is associated with alterations of the ECM, and fibrosis.

  10. Immunoblottings Total collagen was unchanged but the ratio of undenatured collagens to total soluble collagens was increased. Expression and activity of MMPs in heart failure (6). The human failing heart. Support with left ventricular assist devices downregulates matrix metalloproteases, increases TIMPs and reduces collagen damage. Li et al. Circulation 2001, 104: 1152

  11. Infarcted MMP-9 KO mouse Infarcted control mouse Studies with knocked out animal models (1). MMP-9 KO mouse In the mouse, targeted deletion of cardiac MMP-9attenuates left ventricular enlargement and collagen accumulation after experimental infarction.

  12. At 6 weeks, increase in type 3 collagen content. At 6 months, pathological hypertrophy with disruption of the sarcomeric architecture. However, the left ventricular pressure is increased. At 12 months, the contractile function is deteriorated. Control heart Transgenic MMP-1 heart Studies with transgenic animal models (2). Transgenic MMP-1 mouse A mouse that constituvely expresses human MMP-1 Kim et al. JCI, 2000, 106:857 Direct disruption of the extracellular matrix, induced by the overexpression of MMP-1, reproduces changes observed in the progression of human heart failure: initial adaptative response followed by progressive loss of function.

  13. The possible role of MMPs in the development of cardiac fibrosis 1.MMPs are increased in late-stage experimental and human heart failure. 2.TIMPs are decreased in the failing human heart 3. A paradoxal end-result is increased MMPs accompanied with increased fibrosis. Cardiac MMPs not only ensure degradation of matrix components, but also modulate collagen synthesis.: digestion of matrix components leads to the release of biological factors (TGFß, IGF, FGF, TNFalpha..) from the extracellular matrix which initiate neo-synthesis of collagens. Cardiac fibrosis is not only anincrease in matrix collagen, but also changes in collagen type, organization and cross-links.

  14. PD-166793 (Park Davis/ Pfizer) Rapid pacing (S)-2-(4’-bromo-biphenyl-4-sulfonylamino- 3-methyl butyric acid) does not inhibit TACE contrarily to Batimastat and Marimastat. Pharmacological studies (1) Effect of PD166793 in the pig exposed to rapid pacing Spinale et al. Circ. Res. 1999, 85: 364

  15. Parameters Effect of PD-166793 Ventricular remodeling Contraction Collagen content Attenuation Improvement Decrease Pharmacological studies (2). Effect of PD166793 in the rat with progressive heart failure. Peterson et al. Circulation 2001, 103:2303 PD-166793 administration was initiated in 9 month-old rats and continued for 4 months. PD-166793 treatment attenuates the degree of left ventricular remodeling in Spontaneously Hypersentive Heart Failure (SHHF).

  16. CP-471,474 (Pfizer) A broad spectrum inhibitor Echocardiography in the infarcted mouse Pharmacological studies (3) Effect of CP-471,474 in the infarcted mouse Rohde et al. Circulation 1999, 99:3063

  17. Pharmacological studies (4) Effect of CP-471,474 in the infarcted rabbit Lindsey et al. Circulation 2002, 105:753 CP-471,474 (Pfizer) a broad spectrum inhibitor but that does not inhibit MMP-1 in rabbits which like humans, but not rodents, express MMP-1 as a major collagenase.

  18. Our objectives in the RTN project • 1. Definition of experimental animal models of heart failure that display characteristics of the human disease, i.e., activation of MMP-3 and 9 and fibrosis. • - Dahl salt sensitive rats and L-NAME rats, two animal models of heart failure associated with hypertension; • - Infarcted rats. 2. Measurements of cardiac MMP activity and abundance in cardiac tissue fractions in the animals treated or not with MMP inhibitor; - Fluorescent substrates and zymography; - Immunoblotting. • 3. Measurements of functional parameters and disease markers in the animals treated or not with MMP inhibitor • - follow up of the cardiac function parameters by echocardiography; • - collagen cleavage assessed in tissue sections by immunostaining; • - oxidative stress assessed in tissue sections by immunostaining; • - studies of calcium cycle and contraction in isolated, electrically stimulated cardiomyocytes.

  19. - Echocardiographic parameters LVW/ BW d PWLV FS (mg/g) (mm) (%) DR rats (11 w) 3,5 ± 0,3 43,0 ± 0,9 0,09 ± 0,01 DS rats (11 w) 3,5 ± 0,1 0,13 ± 0,01* 50,0 ± 1,4* DR rats (15 w) 3,3 ± 0,1 0,09 ± 0,01 42,9 ± 1,7 DS rats (15 w) 4,1 ± 0,1* 0,11 ± 0,01 39,8 ± 2,3† - Survival 1 0.5 Animal Survival DR rats DS rats 0 10 12 14 Weeks The Dahl salt sensitive rat model (1) Functional parameters - Blood Pressure: 220 mmHg in the Dahl salt sensitive (DS) rat as young as 11 weeks, to compare with 152 mmHg in the Dahl salt resistant (DR) rat.

  20. 11 weeks Collagen staining with red sirius DS 11 weeks DR 11 weeks 15 weeks DR DS Endogenous MMP activity 16 Collagénase activity (nU) 8 0 6 11 15 Age of the rats (weeks) MMP-3 DS rats DR rats Immunoblottings The Dahl salt sensitive rat model (2) Fibrosis and MMPs

  21. Isolated cardiomyocytes loaded with Fura-2 , electrically stimulated and exposed to a positive inotropic agent Ca transients 0.75 F360 / F380 0.55 1200 1260 600 660 0 60 1800 1860 Contraction 8 FS % 0 0 60 1200 1260 600 660 1800 1860 Time (s) Isolated adult rat cardiomyocytes Immunostaining with phalloidin

  22. Participants Sylviane Adubeiro Marie Claude Bourin Michel Cailleret Gabriele Candiani Nicole Defer Catherine Pavoine Françoise Pecker

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