Original Article
MMPs 2 and 9 are essential for coronary collateral growth and are prominently regulated by p38 MAPK

https://doi.org/10.1016/j.yjmcc.2011.08.012Get rights and content

Abstract

Transient, repetitive ischemia (RI) stimulates coronary collateral growth (CCG) in normal, healthy (SD) rats, which requires p38 MAPK activation. In contrast, RI does not induce CCG in the metabolic syndrome (JCR) rats, which is associated with lack of p38 MAPK activation. The functional consequences of p38 MAPK activation in CCG remain unknown. Theoretically, effective collateral growth would require extracellular matrix remodeling; however, direct assessment as well as identification of proteases responsible for this degradation are lacking. In this study, we investigated the role of p38 MAPK in the regulation of matrix metalloproteinases 2 and 9 (MMPs 2 and 9) and their requirement for CCG in SD vs. JCR rats. The rats underwent the RI protocol (8 LAD occlusions, 40 s each, every 20 min, in 8 h cycles for 0, 3, 6, or 9 days). MMP expression was measured in the ischemic, collateral-dependent zone (CZ) and the normal zone (NZ) by Western blot, and MMP activity by zymography. Expression and activation of MMP 2 and 9 were significantly increased (~ 3.5 fold) on day 3 of RI in the CZ of SD rats. In vivo p38 MAPK inhibition completely blocked RI-induced MMP 2 and 9 expression and activation. MMP activation correlated with increased degradation of components of the basement membrane and the vascular elastic laminae: elastin (~ 3 fold), laminin (~ 3 fold) and type IV collagen (~ 2 fold). This was blocked by MMP 2 and 9 inhibition, which also abolished RI-induced CCG. In contrast, in JCR rats, RI did not induce expression or activation of MMP 2 or 9 and there was no associated degradation of elastin, laminin or type IV collagen. In conclusion, MMP 2 and 9 activation is essential for CCG and is mediated, in part, by p38 MAPK. Furthermore, compromised CCG in the metabolic syndrome may be partially due to the lack of p38 MAPK-dependent activation of MMP 2 and 9 and resultant decreased extracellular matrix degradation.

Highlights

► p38 MAPK is required for MMP 2/9 activity during coronary collateral growth (CG). ► MMP 2/9 activity is essential for extracellular matrix (ECM) degradation and CG. ► Lack of CG in metabolic syndrome correlates with lack of MMP 2/9 activity and reduced ECM degradation.

Introduction

Myocardial ischemia–reperfusion injury is a well-known phenomenon resulting from prolonged periods of ischemia followed by re-oxygenation. However, short, repetitive periods of ischemia followed by reperfusion can lead to adaptive responses and render the myocardium tolerant to longer periods of ischemia, in part through promoting collateral development [1]. Stable angina pectoris is a consequence of significant coronary artery constriction, and is characterized by transient periods of ischemia, upon increased myocardial metabolic demand followed by reperfusion at rest. Coronary collateral growth (CCG) is an adaptive response to transient, repetitive myocardial ischemia (RI). Clinically, patients with stable angina have a decreased incidence of fatal myocardial infarction, which is associated with better developed collateral networks [2]. In contrast, CCG has been shown to be severely impaired in patients suffering from type II diabetes [3] and the metabolic syndrome [4]. Likewise, CCG is impaired in our metabolic syndrome rat model (JCR:LA-cp or JCR) [5]. The JCR rat is obese, dyslipidemic (low HDL, high LDL, VLDL, and triglycerides) [5], insulin resistant with impaired glucose tolerance [6], and hypertensive [5], and thus, mimics the complex pathology of the human metabolic syndrome.

The process of CCG involves endothelial and vascular smooth muscle cell (VSMC) proliferation and migration, as well as extracellular matrix (ECM) remodeling. The early phase of collateral growth is associated with inward remodeling, in which cells migrate across the internal elastic lamina and the basement membrane, into the lumen of the pre-existing native collaterals. This is followed by outward remodeling in which cells migrate across the external elastic lamina into the vascular adventitia and the surrounding myocardium, thus allowing for vessel expansion and significant increases in blood flow [7], [8], [9]. Consequently, reorganization of the ECM, including ECM degradation, is a presumed integral part of collateral remodeling. However, direct measurements of this process during collateral growth have never been reported.

ECM degradation requires matrix metalloproteinases (MMPs), zinc-dependent endopeptidases capable of degrading extracellular matrix proteins. MMPs can be separated based on substrate specificity into interstitial collageneases (MMPs 1, 8 and 13), broad specificity MMPs (MMPs 3 and 7), metalloelastases (MMP 12), membrane-bound MMPs (MMP 14 (MT1-MMP) and MMP 17), and gelatinases (MMP 2 and 9). MMP 2 and 9 have been shown to degrade type IV collagen, laminin and elastin, the primary components of the vascular basement membrane and the internal and external elastic laminae, in vitro [10], [11], [12], [13]. They are known to play a role in cell proliferation, migration, differentiation, angiogenesis associated with cancer metasthesis, neointima formation following vascular injury and aneurysm formation and rupture [14], [15], [16]. Although degradation of the basement membrane and the vascular elastic laminae is a common aspect shared between these processes and collateral remodeling, they are not identical, and conclusions drawn from these studies do not uniformly apply to collateral growth. Increased MMP 2 and 9 expression has been associated with collateral growth, but the results are not entirely in agreement. In one study, during the early, inward remodeling phase in growing coronary collaterals, the neointima showed high expression of MMPs 2 and 9 while mature collaterals expressed low levels of these MMPs [17]. On the other hand, MMP 2 but not MMP 9 expression and activity were increased in mesenteric collateral vessels [18]. Importantly, a conclusive requirement for MMP 2 and 9 activation in CCG has not been shown. Furthermore, it is unknown whether MMP 2 and/or 9 regulation is altered in the metabolic syndrome, where CCG is impaired.

MMPs are regulated at the level of both expression and activation. Several signaling pathways have been shown to regulate MMP expression and/or activation. Among these are the mitogen activated protein kinases (MAPKs). MAPKs can be divided into the extracellular signal-regulated kinases (ERK1/2), p38 MAPK, and c-Jun N-terminal kinase (JNK) [19]. The ERK1/2 pathway has been shown to regulate expression and activation of many MMPs including MMP 9 [20]. p38 MAPK, when activated during inflammation or the innate immune response, lead to the activation of MMP 9 [21], and a single study implicates p38 MAPK in the regulation of MMP 9 activation in cultured airway smooth muscle cells [22]. However, it is not known which signaling pathways may regulate MMP expression and/or activation in collateral growth.

We have previously shown that transient p38 MAPK activation on day 3 of RI is required for CCG in the normal healthy rat model where inhibition of p38 MAPK resulted in ~ 60% reduction in RI-induced CCG [23]. In addition, we have shown that RI-induced CCG was severely compromised in the metabolic syndrome JCR rat model, and that this correlated with lack of RI-induced p38 MAPK activation [24]. However, the functional consequence of p38 MAPK activation in collateral growth remained unknown. Therefore the goals of this study were to determine: 1) whether RI-induced activation of p38 MAPK regulated the development of coronary collaterals through the activation of MMP 2 and 9 and the degradation of their ECM substrates, 2) whether MMP 2 and 9 were required for this ECM degradation and CCG, and 3) whether MMP 2 and/or 9 expression and/or activity were altered in the metabolic syndrome.

Section snippets

Rat model of coronary collateral growth/RI

Male, 10–12 week old Sprague–Dawley (SD) (300–350 g) or obese JCR:LA-cp rats (JCR) (650–700 g) were used for chronic implantation of a pneumatic occluder over the left anterior descending coronary artery (LAD), as described previously [25]. The RI protocol for rats consisted of 8 40s occlusions, one every 20 min over 2 h 20 min followed by a period of “rest” for 5 h 40 min. This 8 h cycle was repeated three times per day for 0–9 days. The specific inhibitors of p38 MAPK, SB203580 (3.2 mg/kg/day,

MMP 2 and 9 expression and activation are increased in response to repetitive ischemia in normal, healthy animals but not in the metabolic syndrome

No significant basal expression or activation of either MMP2 or 9 was observed in either rat phenotype. Western blot analysis demonstrated an increase in expression of MMP 2 and 9 in the SD animals specifically confined to the collateral dependent zone (CZ) on day 3 of the RI protocol (3.3 ± 0.3 fold vs. NZ for MMP 2; 3.6 ± 0.2 fold vs. NZ for MMP 9) (Fig. 1A). This increase in expression in the collateral dependent zone corroborates our previously published data that showed increased p38 MAPK

Discussion

The major novel findings in this study are that: 1) RI induces the expression and activation of MMP 2 and 9, which correlates with degradation of components of the basement membrane and the elastic laminae, type IV collagen, laminin and elastin, 2) RI-induced activation of MMP 2 and 9 is severely compromised in the metabolic syndrome, which is associated with the absence of type IV collagen, laminin and elastin degradation, 3) MMP 2 and 9 expression and activation by RI as well as type IV

Disclosure statement

None declared.

Acknowledgment

This study was supported by NIH R01 HL093052.

Glossary

CCG
coronary collateral growth
CZ
collateral-dependent zone, LAD perfusion territory
ECM
extracellular matrix
ERK1/2 MAPK
extracellular signal-regulated mitogen-activated protein kinase
FGF
fibroblast growth factor
JNK MAPK
c-Jun mitogen-activated protein kinase
JCR
Russell rat, JCR:LA-cp
LAD
left anterior descending coronary artery
MK2
mitogen-activated protein kinase activated protein kinase 2
MMP
matrix metalloproteinase
NZ
normal, non-ischemic zone
p38 MAPK
p38 mitogen-activated protein kinase
RI
repetitive ischemia

References (79)

  • V. Lagente et al.

    Role of matrix metalloproteinases in the inflammatory process of respiratory diseases

    J Mol Cell Cardiol

    (2010)
  • S. Rousseau et al.

    Integrating the VEGF signals leading to actin-based motility in vascular endothelial cells

    Trends Cardiovasc Med

    (2000)
  • A. Cuenda et al.

    p38 MAP-kinases pathway regulation, function and role in human diseases

    Biochim Biophys Acta

    (2007)
  • S. Luangdilok et al.

    MAPK and PI3K signalling differentially regulate angiogenic and lymphangiogenic cytokine secretion in squamous cell carcinoma of the head and neck

    Eur J Cancer

    (2011)
  • N. Hiraoka et al.

    Matrix metalloproteinases regulate neovascularization by acting as pericellular fibrinolysins

    Cell

    (1998)
  • C. Steingen et al.

    Characterization of key mechanisms in transmigration and invasion of mesenchymal stem cells

    J Mol Cell Cardiol

    (2008)
  • W. Chen et al.

    A redox-based mechanism for cardioprotection induced by ischemic preconditioning in perfused rat heart

    Circ Res

    (1995)
  • C. Seiler

    The human coronary collateral circulation

    Heart

    (2003)
  • J. Waltenberger

    Impaired collateral vessel development in diabetes: potential cellular mechanisms and therapeutic implications

    Cardiovasc Res

    (2001)
  • M.B. Yilmaz et al.

    Obesity is associated with impaired coronary collateral vessel development

    Int J Obes Relat Metab Disord

    (2003)
  • R. Reed et al.

    The mechanistic basis for the disparate effects of angiotensin II on coronary collateral growth

    Arterioscler Thromb Vasc Biol

    (2008)
  • D. Wilson et al.

    Low matrix metalloproteinase levels precede vascular lesion formation in the JCR:LA-cp rat

    Mol Cell Biochem

    (2003)
  • D. Scholz et al.

    Arteriogenesis, a new concept of vascular adaptation in occlusive disease

    Angiogenesis

    (2001)
  • J.L. Unthank et al.

    Early adaptations in collateral and microvascular resistances after ligation of the rat femoral artery

    J Appl Physiol

    (1995)
  • W. Schaper

    Collateral circulation: past and present

    Basic Res Cardiol

    (2009)
  • C.B. Jones et al.

    Matrix metalloproteinases: a review of their structure and role in acute coronary syndrome

    Cardiovasc Res

    (2003)
  • G. Giannelli et al.

    Induction of cell migration by matrix metalloprotease-2 cleavage of laminin-5

    Science

    (1997)
  • P.E. Van den Steen et al.

    Biochemistry and molecular biology of gelatinase B or matrix metalloproteinase-9 (MMP-9)

    Crit Rev Biochem Mol Biol

    (2002)
  • C.M. Overall et al.

    Strategies for MMP inhibition in cancer: innovations for the post-trial era

    Nat Rev Cancer

    (2002)
  • W.J. Cai et al.

    Remodeling of the vascular tunica media is essential for development of collateral vessels in the canine heart

    Mol Cell Biochem

    (2004)
  • T.L. Haas et al.

    Involvement of MMPs in the outward remodeling of collateral mesenteric arteries

    Am J Physiol Heart Circ Physiol

    (2007)
  • M.P. Vincenti et al.

    Signal transduction and cell-type specific regulation of matrix metalloproteinase gene expression: can MMPs be good for you?

    J Cell Physiol

    (2007)
  • W. Eberhardt et al.

    Amplification of IL-1 beta-induced matrix metalloproteinase-9 expression by superoxide in rat glomerular mesangial cells is mediated by increased activities of NF-kappa B and activating protein-1 and involves activation of the mitogen-activated protein kinase pathways

    J Immunol

    (2000)
  • K.C. Liang et al.

    Interleukin-1beta induces MMP-9 expression via p42/p44 MAPK, p38 MAPK, JNK, and nuclear factor-kappaB signaling pathways in human tracheal smooth muscle cells

    J Cell Physiol

    (2007)
  • P. Rocic et al.

    Optimal reactive oxygen species concentration and p38 MAP kinase are required for coronary collateral growth

    Am J Physiol Heart Circ Physiol

    (2007)
  • R. Reed et al.

    Redox-sensitive Akt and Src regulate coronary collateral growth in metabolic syndrome

    Am J Physiol Heart Circ Physiol

    (2009)
  • E. Toyota et al.

    Vascular endothelial growth factor is required for coronary collateral growth in the rat

    Circulation

    (2005)
  • R. Jadhav et al.

    Angiotensin type I receptor blockade in conjunction with enhanced Akt activation restores coronary collateral growth in the metabolic syndrome

    Am J Physiol Heart Circ Physiol

    (2011)
  • S. Monaco et al.

    Enzymatic processing of collagen IV by MMP-2 (gelatinase A) affects neutrophil migration and it is modulated by extracatalytic domains

    Protein Sci

    (2006)
  • Cited by (41)

    • Timosaponin AIII: A novel potential anti-tumor compound from Anemarrhena asphodeloides

      2018, Steroids
      Citation Excerpt :

      Several major mitogen-activated protein kinases (MAPKs) signaling pathways were related with cell migration and invasion including extracellular signal regulated kinase 1 and (ERK1/2), p38 and c-Jun N-terminal kinase (JNK). They have been also proved that be involved in the activations of various MMPs [36]. In A549 cells treated by timosaponin AIII, the activation of ERK1/2 was significantly inhibited while activations of p38 and JNK were not affected.

    • MMP-2 and MMP-9 contribute to the angiogenic effect produced by hypoxia/15-HETE in pulmonary endothelial cells

      2018, Journal of Molecular and Cellular Cardiology
      Citation Excerpt :

      So these findings indicate that MMP-2 and MMP-9 are the downstream targets of 15-LO/15-HETE pathway in vivo and in vitro. Previous studies have reported that the p38 MAPK signaling is involved in MMP-2 and MMP-9 activation [28]. We detected the expression of p-p38, p38 in NDGA treated cell.

    • Elevated 20-HETE in metabolic syndrome regulates arterial stiffness and systolic hypertension via MMP12 activation

      2018, Journal of Molecular and Cellular Cardiology
      Citation Excerpt :

      In our study, MMP2, MMP9, MMP3 or MMP7 are not involved in 20-HETE-dependent regulation of elastin degradation or regulation of arterial stiffness. This is consistent with our previous observations that activation of these MMPs is decreased in arteries from metabolic syndrome animals [48,89]. However, other elastases, for example cathepsin G from neutrophils, may play a role in elastin degradation.

    • Lentil consumption reduces resistance artery remodeling and restores arterial compliance in the spontaneously hypertensive rats

      2016, Journal of Nutritional Biochemistry
      Citation Excerpt :

      Rather, the apparent but nonsignificant decline in collagen content relative to the nsECM of the arterial wall suggests that collagen is being replaced by another extracellular component. Given the close connection between p38MAPK and ECM production and degradation [36,39,40], this component may be a distinct ECM protein, matrix-metalloproteinase-degraded collagen or a combination of both, but at this point, its identity remains unknown. As well, it is possible that the nsECM consists of glycosaminoglycan, which has been shown to be increased in the aorta of SHR and decreased by a diet containing high levels of wild blueberry [41].

    View all citing articles on Scopus
    View full text