TIMP2 Deficiency Accelerates Adverse Post–Myocardial Infarction Remodeling Because of Enhanced MT1-MMP Activity Despite Lack of MMP2 Activation

Abstract

Rationale: Myocardial infarction (MI) results in remodeling of the myocardium and the extracellular matrix (ECM). Tissue inhibitors of metalloproteinases (TIMPs) are critical regulators of ECM integrity via inhibiting matrix metalloproteinases (MMPs). TIMP2 is highly expressed in the heart and is the only TIMP that, in addition to inhibiting MMPs, is required for cell surface activation of pro-MMP2. Hence, it is difficult to predict the function of TIMP2 as protective (MMP-inhibiting) or harmful (MMP-activating) in heart disease. Objective: We examined the role of TIMP2 in the cardiac response to MI. Methods and Results: MI was induced in 11-to 12-week-old male TIMP2؊/؊ and age-matched wild-type mice. Cardiac function was monitored by echocardiography at 1 and 4 weeks post-MI. ECM fibrillar structure was visualized using second harmonic generation and multiphoton imaging of unfixed/unstained hearts. Molecular analyses were performed at 3 days and 1 week post-MI on flash-frozen infarct, periinfarct, and noninfarct tissue. Membrane type 1 (MT1)-MMP levels and activity were measured in membrane protein fractions. TIMP2؊/؊ -MI mice exhibited a 25% greater infarct expansion, markedly exacerbated left ventricular dilation (by 12%) and dysfunction (by 30%), and more severe inflammation compared to wild-type MI mice. Adverse ECM remodeling was detected by reduced density and enhanced disarray of fibrillar collagen in TIMP2 ؊/؊ -MI compared to wild-type MI hearts. TIMP2 deficiency completely abrogated MMP2 activation but markedly increased collagenase activity, particularly MT1-MMP activity post-MI. Key Words: myocardial infarction Ⅲ extracellular matrix Ⅲ remodeling Ⅲ tissue inhibitor of metalloproteinase Ⅲ matrix metalloproteinase C oronary artery disease is the most common cause of heart failure in Western nations. 1 Myocardial infarction (MI) results in adverse remodeling of the myocardium including the extracellular matrix (ECM). 2 ECM is a dynamic structure which undergoes constant turnover, and its intact structure is essential for optimal cardiac structure and function. Tissue inhibitors of metalloproteinases (TIMPs) play a critical role in regulating the activity of the ECM-degrading enzymes matrix metalloproteinases (MMPs). [3][4][5] Four TIMPs have been identified so far (TIMP1 to TIMP4), with TIMP2 and TIMP3 being the highly expressed TIMPs in the heart and TIMP4 showing a tissue-specific expression pattern which includes the heart and the brain. 6 TIMP2 levels have been shown to be altered in patients with different types of heart disease such as pressure overload 7 and atrial fibrillation. 8 In patients with ischemic or idiopathic dilated cardiomyopathy, myocardial protein levels of TIMP2 remained unaltered 9,10 or increased in end-stage dilated cardiomyopathy patients, 11 whereas plasma analyses revealed a rise in TIMP2 levels in late stages of MI. 12 TIMP2 is unique among TIMPs because, in addition to inhibiting a number of MMPs, it mediates MMP activation. TIMP2 is required for cell surface activati...

Age- and gender-dependent myocardial transcription patterns of cytokines and extracellular matrix remodelling enzymes in cats with non-cardiac diseases

“…In mouse models, Th2 induction resulted in reduced ventricular stiffness, and Th1 cells were involved in initiation of fibrosis and collagen crosslinking (Yu et al, 2005;, whereasanother 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 21 study did detecta shift from Th1 to Th2 response in old mice in association with interstitial myocardial fibrosis (Cieslik et al, 2011). We observed a reduction of all three immune modulatory markers (IL-2, IL-4,IFN-γ)with age in thefemale cats, which might be consistent with a generally reduced myocardial inflammatory response, as suggested by thereduced transcriptionof IL-1 and IL-6.Aupperle and co-authors (2011) observed reduced TIMP-2 expression in association with myocardial fibrosis in cats.In older sheep (> 8 years), rat and mouse models, higher myocardial MMP-2 and reduced TIMP-2 mRNA and protein expression, as well as cardiomyocyte hypertrophy and perivascular and interstitial fibrosis, imbalanced cardiac remodelling and impaired repair was observed (Kandalam et al, 2010;Wang et al, 2010;Horn et al, 2012;Givvimani et al, 2013). The age-associated reduced TIMP-2 and MMP-2 transcription in female and male cats, respectively,suggests therefore a pro-fibrotic potential and impaired repair of the female myocardium, and further supports a reactive male myocardium facilitating appropriate repair (Davis et al, 2007;Chen and Frangogiannis, 2010).…”

“…Matrix metalloproteinase and TIMP imbalance and ECM degradation and deposition are associated withcardiac diseases and dysfunction (Spinale, 2007), but onlylittle is known about their role in age related cardiac changes. Mouse models ofageingexhibit varying MMP, TIMP and transforming growth factor (TGF)-β mRNA and protein production (Brooks and Conrad, 2000;Lindsey et al, 2005;Tian et al, 2007;Kandalam et al, 2010;Wang et al, 2010;Givvimani et al, 2013;Uchinaka et al, 2014). 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 5 Cytokines play an important role in the activation of MMPs and TIMPs.…”

Loss of Timp3 Gene Leads to Abdominal Aortic Aneurysm Formation in Response to Angiotensin II

“…Western blotting was performed to detect collagen type I (COL1A1) (Santa Cruz Biotechnology) and ␣-elastin (Abcam Inc.), and in vitro gelatin zymography was carried out as before (30). A Coomassie Bluestained SDS-PAGE gel (after proteins were transferred to PVDF membrane) was used as a loading control.…”

“…In situ gelatin zymography was performed on 14-m cryosections as described (31). Total collagenase and gelatinase activities were measured using Enzchek collagenase (catalog number D-12060) and elastase (catalog number E-12056) activity assay kits (from Millipore) as before (30). Briefly, 50 g of protein extract was incubated with fluorescently labeled collagen or elastin substrate.…”

“…The generated fluorescence signal was recorded for 4 h (excitation, 485 nm; emission, 515 nm), and the slopes of the curves were used to calculate the collagenase or elastase activities. mRNA expression was measured using TaqMan real time PCR (Table 1) (30,32).…”