The role of matrix metalloproteinases induced angiogenesis during transition from compensatory cardiac hypertrophy to cardiac failure : therapeutic effects of hydrogen sulfide.

Background: Although matrix metalloproteinase (MMPs) and tissue inhibitor of metalloproteinase (TIMPs) play a vital role in tumor angiogenesis and TIMP-3 causes apoptosis, their role in cardiac angiogenesis is unknown. Interestingly, a disruption of coordinated cardiac hypertrophy and angiogenesis contributes to the transition to heart failure, however, the proteolytic and anti-angiogenic mechanisms of transition from compensatory hypertrophy to decompensatory heart failure are unclear. Previous studies have shown the cardio protective role of hydrogen sulfide (H2S) in myocardial ischemia, infarction and heart failure, but its role during transition from compensatory cardiac hypertrophy to heart failure is yet to be unveiled. We hypothesize that in pathological pressure overload of the heart, cardiac matrix remodeling is induced by an increase in angiogenic growth factors during the compensatory hypertrophy phase and an increase in antiangiogenic factors during the decompensatory heart failure phase. H2S mitigates the transition from compensatory hypertrophy to decompensatory heart failure by increasing angiogenesis and decreasing fibrosis. Methods: In the first set of experiments, we have created ascending aortic banding (AB) in the mice to mimic pressure overload on the heart and studied the ventricular remodeling events associated with chronic pressure overload. Male wild type mice were used and all animal procedures were performed in accordance with National Institute of Health guidelines for animal research and were reviewed and approved by the Institute Animal Care and use Committee of University of Louisville. Sham animals underwent similar procedures except for aortic banding. Animals were studied at 3 weeks (compensatory phase) and 8 weeks period (decompensatory phase) and compared with sham groups. In the second set of experiment, similar protocol was followed, and animals were treated with hydrogen sulfide (H2S) for 6 weeks by giving sodium hydrosulfide (NaHS) in drinking water and compared with untreated groups. Molecular and functional data were assessed by echocardiography, pressure-volume (P-V) study, immunohistochemistry, histology, western blot, and x-ray angiography. Results: We found that in the first set of experiments, expression of MMP-2 increased along with angiogenic growth factor, vascular endothelial growth factor (VEGF) during compensatory phase (AB 3 weeks group). Expression of MMP-9, TIMP-3 and anti-angiogenic factors, angiostatin, and endostatin increased during decompensatory phase (AB 8 weeks group). There was increased deposition of fibrosis during the decompensatory phase. Sy treating with H2S, we noticed that there was increased expression of VEGF and MMP-2 in AS 8 weeks group than in untreated AS 8 weeks group. Interestingly, the expression of MMP-9, TIMP-3 and anti-angiogenic factors angiostatin and endostatin decreased in H2S treated AS 8 weeks group than in the untreated group. There was significant reduction of fibrosis in treated AS 8 weeks group. Left ventricular function also improved in H2S treated AS 8 weeks group. Conclusion: we conclude that under chronic pressure overload, expression of MMP-2 is induced which in turn increases the release of angiogenic growth factors in compensatory cardiac hypertrophy. The expression of MMP-9 and TIMP-3 is also increased leading to a release of anti-angiogenic factors in decompensatory heart failure. Hydrogen sulfide mitigates this transition to decompensatory heart failure not only by inducing MMP-2NEGF but also inhibiting MMP-9/TIMP-3 and anti-angiogenic factors

Mitochondrial division/mitophagy inhibitor (Mdivi) Ameliorates Pressure Overload Induced Heart Failure

Background: We have previously reported the role of anti-angiogenic factors in inducing the transition from compensatory cardiac hypertrophy to heart failure and the significance of MMP-9 and TIMP-3 in promoting this process during pressure overload hemodynamic stress. Several studies reported the evidence of cardiac autophagy, involving removal of cellular organelles like mitochondria (mitophagy), peroxisomes etc., in the pathogenesis of heart failure. However, little is known regarding the therapeutic role of mitochondrial division inhibitor (Mdivi) in the pressure overload induced heart failure. We hypothesize that treatment with mitochondrial division inhibitor (Mdivi) inhibits abnormal mitophagy in a pressure overload heart and thus ameliorates heart failure condition. Materials and Methods: To verify this, ascending aortic banding was done in wild type mice to create pressure overload induced heart failure and then treated with Mdivi and compared with vehicle treated controls. Results: Expression of MMP-2, vascular endothelial growth factor, CD31, was increased, while expression of anti angiogenic factors like endostatin and angiostatin along with MMP-9, TIMP-3 was reduced in Mdivi treated AB 8 weeks mice compared to vehicle treated controls. Expression of mitophagy markers like LC3 and p62 was decreased in Mdivi treated mice compared to controls. Cardiac functional status assessed by echocardiography showed improvement and there is also a decrease in the deposition of fibrosis in Mdivi treated mice compared to controls

Cardiac matrix: A clue for future therapy

AbstractCardiac muscle is unique because it contracts ceaselessly throughout the life and is highly resistant to fatigue. The marvelous nature of the cardiac muscle is attributed to its matrix that maintains structural and functional integrity and provides ambient micro-environment required for mechanical, cellular and molecular activities in the heart. Cardiac matrix dictates the endothelium myocyte (EM) coupling and contractility of cardiomyocytes. The matrix metalloproteinases (MMPs) and their tissue inhibitor of metalloproteinases (TIMPs) regulate matrix degradation that determines cardiac fibrosis and myocardial performance. We have shown that MMP-9 regulates differential expression of micro RNAs (miRNAs), calcium cycling and contractility of cardiomyocytes. The differential expression of miRNAs is associated with angiogenesis, hypertrophy and fibrosis in the heart. MMP-9, which is involved in the degradation of cardiac matrix and induction of fibrosis, is also implicated in inhibition of survival and differentiation of cardiac stem cells (CSC). Cardiac matrix is distinct because it renders mechanical properties and provides a framework essential for differentiation of cardiac progenitor cells (CPC) into specific lineage. Cardiac matrix regulates myocyte contractility by EM coupling and calcium transients and also directs miRNAs required for precise regulation of continuous and synchronized beating of cardiomyocytes that is indispensible for survival. Alteration in the matrix homeostasis due to induction of MMPs, altered expression of specific miRNAs or impaired signaling for contractility of cardiomyocytes leads to catastrophic effects. This review describes the mechanisms by which cardiac matrix regulates myocardial performance and suggests future directions for the development of treatment strategies in cardiovascular diseases

Activated caspace-3 staining of mouse cardiac sections showing apoptosis in sham, 8 weeks post-AB (AB 8 wks) treated with vehicle control and Mdivi.

<p><b>A</b>) Apoptotic cells are seen as red fluorescent dots (scale bar- 20 µm). <b>C</b>) Activated caspace-3 +ve cells represented in an enlarged area. <b>B</b>) Data represents mean ±SE from n = 6 per group; *p<0.05 was considered significant compared to sham and ^#p<0.05 compared to vehicle treated group.</p

Folic acid mitigated cardiac dysfunction by normalizing the levels of tissue inhibitor of metalloproteinase and homocysteine-metabolizing enzymes postmyocardial infarction in mice

Myocardial infarction (MI) results in significant metabolic derangement, causing accumulation of metabolic by product, such as homocysteine (Hcy). Hcy is a nonprotein amino acid generated during nucleic acid methylation and demethylation of methionine. Folic acid (FA) decreases Hcy levels by remethylating the Hcy to methionine, by 5-methylene tetrahydrofolate reductase (5-MTHFR). Although clinical trials were inconclusive regarding the role of Hcy in MI, in animal models, the levels of 5-MTHFR were decreased, and FA mitigated the MI injury. We hypothesized that FA mitigated MI-induced injury, in part, by mitigating cardiac remodeling during chronic heart failure. Thus, MI was induced in 12-wk-old male C57BL/J mice by ligating the left anterior descending artery, and FA (0.03 g/l in drinking water) was administered for 4 wk after the surgery. Cardiac function was assessed by echocardiography and by a Millar pressure-volume catheter. The levels of Hcy-metabolizing enzymes, cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 5-MTHFR, were estimated by Western blot analyses. The results suggest that FA administered post-MI significantly improved cardiac ejection fraction and induced tissue inhibitor of metalloproteinase, CBS, CSE, and 5-MTHFR. We showed that FA supplementation resulted in significant improvement of myocardial function after MI. The study eluted the importance of homocysteine (Hcy) metabolism and FA supplementation in cardiovascular disease

IHC staining of mouse heart sections with mitochondrial marker antibody MTCO-2 in sham, 8 weeks post-AB (AB 8 wks) treated with vehicle control and Mdivi.

<p><b>A</b>) Expression of MTCO-2 is seen as red fluorescence intensity (scale bar-50 µm). <b>B</b>) Data represents mean ±SE from n = 6 per group; *p<0.05 was considered significant compared to sham and ^#p<0.05 compared to vehicle treated group.</p