Protective role of polyphenols in cyclosporine A-induced nephrotoxicity during rat pregnancy.

The aim of this study was to evaluate the adverse effects of cyclosporine A (CsA) toward renal morphogenesis and to test the renoprotective natural antioxidants such as provinol (PV). Pregnant rats were divided into four groups. Group I was injected SC with olive oil. Group II was treated with oral administration of PV and was used as control. Group III animals were injected SC daily with CsA, and group IV animals were injected daily with CsA and PV for 21 days of pregnancy. Five pups per litter were killed and the kidneys removed and treated by morphological and immunohistochemical (IHC) methods. IHC analysis considered two proteins responsible for nephrotoxicity in adult rats: inducible nitric oxide (iNOS) and matrix metalloproteinase-2 (MMP2). Pregnancy outcomes among CsA-treated rats demonstrated a reduced number of pups. Pups that were exposed antenatally to CsA presented several pathologic findings in all immature parenchyma and an increase in iNOS and MMP2 expression. These side effects were not observed in kidney of litters born from CsA + PV-treated mothers. Our study indicates that CsA induces morphological alterations in renal parenchyma of neonates and that PV plays a protective role against these side effects.     

Temporal and Molecular Analyses of Cardiac Extracellular Matrix Remodeling following Pressure Overload in Adiponectin Deficient Mice

Adiponectin, circulating levels of which are reduced in obesity and diabetes, mediates cardiac extracellular matrix (ECM) remodeling in response to pressure overload (PO). Here, we performed a detailed temporal analysis of progressive cardiac ECM remodelling in adiponectin knockout (AdKO) and wild-type (WT) mice at 3 days and 1, 2, 3 and 4 weeks following the induction of mild PO via minimally invasive transverse aortic banding. We first observed that myocardial adiponectin gene expression was reduced after 4 weeks of PO, whereas increased adiponectin levels were detected in cardiac homogenates at this time despite decreased circulating levels of adiponectin. Scanning electron microscopy and Masson’s trichrome staining showed collagen accumulation increased in response to 2 and 4 weeks of PO in WT mice, while fibrosis in AdKO mice was notably absent after 2 weeks but highly apparent after 4 weeks of PO. Time and intensity of fibroblast appearance after PO was not significantly different between AdKO and WT animals. Gene array analysis indicated that MMP2, TIMP2, collagen 1α1 and collagen 1α3 were induced after 2 weeks of PO in WT but not AdKO mice. After 4 weeks MMP8 was induced in both genotypes, MMP9 only in WT mice and MMP1α only in AdKO mice. Direct stimulation of primary cardiac fibroblasts with adiponectin induced a transient increase in total collagen detected by picrosirius red staining and collagen III levels synthesis, as well as enhanced MMP2 activity detected via gelatin zymography. Adiponectin also enhanced fibroblast migration and attenuated angiotensin-II induced differentiation to a myofibroblast phenotype. In conclusion, these data indicate that increased myocardial bioavailability of adiponectin mediates ECM remodeling following PO and that adiponectin deficiency delays these effects.     

Ablation of MMP9 induces survival and differentiation of cardiac stem cells into cardiomyocytes in the heart of diabetics: a role of extracellular matrix

The contribution of extracellular matrix (ECM) to stem cell survival and differentiation is unequivocal, and matrix metalloproteinase-9 (MMP9) induces ECM turn over; however, the role of MMP9 in the survival and differentiation of cardiac stem cells is unclear. We hypothesize that ablation of MMP9 enhances the survival and differentiation of cardiac stem cells into cardiomyocytes in diabetics. To test our hypothesis, Ins2(+/-) Akita, C57 BL/6J, and double knock out (DKO: Ins2(+/-)/MMP9(-/-)) mice were used. We created the DKO mice by deleting the MMP9 gene from Ins2(+/-). The above 3 groups of mice were genotyped. The activity and expression of MMP9 in the 3 groups were determined by in-gel gelatin zymography, Western blotting, and confocal microscopy. To determine the role of MMP9 in ECM stiffness (fibrosis), we measured collagen deposition in the histological sections of hearts using Masson's trichrome staining. The role of MMP9 in cardiac stem cell survival and differentiation was determined by co-immunoprecipitation (co-IP) of MMP9 with c-kit (a marker of stem cells) and measuring the level of troponin I (a marker of cardiomyocytes) by confocal microscopy in the 3 groups. Our results revealed that ablation of MMP9 (i) reduces the stiffness of ECM by decreasing collagen accumulation (fibrosis), and (ii) enhances the survival (elevated c-kit level) and differentiation of cardiac stem cells into cardiomyocytes (increased troponin I) in diabetes. We conclude that inhibition of MMP9 ameliorates stem cell survival and their differentiation into cardiomyocytes in diabetes.     

Mutations in MMP9 and MMP13 Determine the Mode of Inheritance and the Clinical Spectrum of Metaphyseal Anadysplasia

The matrix metalloproteinases MMP9 and MMP13 catalyze the degradation of extracellular matrix (ECM) components in the growth plate and at the same time cleave and release biologically active molecules stored in the ECM, such as VEGFA. In mice, ablation of Mmp9, Mmp13, or both Mmp9 and Mmp13 causes severe distortion of the metaphyseal growth plate. We report that mutations in either MMP9 or MMP13 are responsible for the human disease metaphyseal anadysplasia (MAD), a heterogeneous group of disorders for which a milder recessive variant and a more severe dominant variant are known. We found that recessive MAD is caused by homozygous loss of function of either MMP9 or MMP13, whereas dominant MAD is associated with missense mutations in the prodomain of MMP13 that determine autoactivation of MMP13 and intracellular degradation of both MMP13 and MMP9, resulting in a double enzymatic deficiency.     

Absence of tissue inhibitor of metalloproteinases 3 disrupts alveologenesis in the mouse

Tissue inhibitors of metalloproteinases (TIMPs) regulate extracellular matrix (ECM) degradation by matrix metalloproteinases (MMPs) throughout lung development. We examined lungs from TIMP3 null mice and found significant air space enlargement compared with wild type (WT) animals during a time course spanning early alveologenesis (post‐partum days 1, 5, 9 and 14). Trichrome staining revealed a similar pattern of collagen distribution in the walls of nascent alveoli; however, the alveolar walls of TIMP3 mutant mice appeared to be thinner than controls. Assessment of MMP2 and MMP9 activities by gelatin zymography demonstrated a significant elevation in the active form of MMP2 at post‐partum days 1 and 5. Treatment of null pregnant dams with a broad spectrum synthetic metalloproteinase inhibitor, GM6001, on embryonic day 16.5 enhanced the formation of primitive alveoli during the saccular stage of lung development as evidenced by a partial, but significant, rescue of alveolar size in post‐partum day 1 animals. We propose that increased MMP activity in the absence of TIMP3 enhances ECM proteolysis, upsetting proper formation of primitive alveolar septa during the saccular stage of alveologenesis. Therefore, TIMP3 indirectly regulates alveolar formation in the mouse. To our knowledge, ours is the first study to demonstrate that in utero manipulation of the TIMP/MMP proteolytic axis, to specifically inhibit proteolysis, significantly affects lung development.     

Blueberry flavonoids inhibit matrix metalloproteinase activity in DU145 human prostate cancer cells.

Regulation of the matrix metalloproteinases (MMPs), the major mediators of extracellular matrix (ECM) degradation, is crucial to regulate ECM proteolysis, which is important in metastasis. This study examined the effects of 3 flavonoid-enriched fractions (a crude fraction, an anthocyanin-enriched fraction, and a proanthocyanidin-enriched fraction), which were prepared from lowbush blueberries (Vaccinium angustifolium), on MMP activity in DU145 human prostate cancer cells in vitro. Using gelatin gel electrophoresis, MMP activity was evaluated from cells after 24-hr exposure to blueberry fractions. All fractions elicited an ability to decrease the activity of MMP-2 and MMP-9. Of the fractions tested, the proanthocyanidin-enriched fraction was found to be the most effective at inhibiting MMP activity in these cells. No induction of either necrotic or apoptotic cell death was noted in these cells in response to treatment with the blueberry fractions. These findings indicate that flavonoids from blueberry possess the ability to effectively decrease MMP activity, which may decrease overall ECM degradation. This ability may be important in controlling tumor metastasis formation.     

Altered endochondral bone development in matrix metalloproteinase 13-deficient mice

The assembly and degradation of extracellular matrix (ECM) molecules are crucial processes during bone development. In this study, we show that ECM remodeling is a critical rate-limiting step in endochondral bone formation. Matrix metalloproteinase (MMP) 13 (collagenase 3) is poised to play a crucial role in bone formation and remodeling because of its expression both in terminal hypertrophic chondrocytes in the growth plate and in osteoblasts. Moreover, a mutation in the human MMP13 gene causes the Missouri variant of spondyloepimetaphyseal dysplasia. Inactivation of Mmp13 in mice through homologous recombination led to abnormal skeletal growth plate development. Chondrocytes differentiated normally but their exit from the growth plate was delayed. The severity of the Mmp13- null growth plate phenotype increased until about 5 weeks and completely resolved by 12 weeks of age. Mmp13-null mice had increased trabecular bone, which persisted for months. Conditional inactivation of Mmp13 in chondrocytes and osteoblasts showed that increases in trabecular bone occur independently of the improper cartilage ECM degradation caused by Mmp13 deficiency in late hypertrophic chondrocytes. Our studies identified the two major components of the cartilage ECM, collagen type II and aggrecan, as in vivo substrates for MMP13. We found that degradation of cartilage collagen and aggrecan is a coordinated process in which MMP13 works synergistically with MMP9. Mice lacking both MMP13 and MMP9 had severely impaired endochondral bone, characterized by diminished ECM remodeling, prolonged chondrocyte survival, delayed vascular recruitment and defective trabecular bone formation (resulting in drastically shortened bones). These data support the hypothesis that proper ECM remodeling is the dominant rate-limiting process for programmed cell death, angiogenesis and osteoblast recruitment during normal skeletal morphogenesis.     

Matrix metalloproteinase stromelysin-3 in development and pathogenesis

The extracellular matrix (ECM) serves as a medium for cell-cell interactions and can directly signal cells through cell surface ECM receptors, such as integrins. In addition, many growth factors and signaling molecules are stored in the ECM. Thus, ECM remodeling and/or degradation plays a critical role in cell fate and behavior during many developmental and pathological processes. ECM remodeling/degradation is, to a large extent, mediated by matrix metalloproteinases (MMPs), a family of extracellular or membrane-bound, Zn2+-dependent proteases that are capable of digesting various proteinaceous components of the ECM. Of particular interest among them is the MMP11 or stromelysin-3, which was first isolated as a breast cancer associated protease. Here, we review some evidence for the involvement of this MMP in development and diseases with a special emphasis on amphibian metamorphosis, a postembryonic, thyroid hormone-dependent process that transforms essentially every organ/tissue of the animal.     

Matrix metalloproteinase-14 is a mechanically regulated activator of secreted MMPs and invasion

Matrix metalloproteinases (MMPs) are extracellular matrix (ECM) degrading enzymes and have complex and specific regulation networks. This includes activation interactions, where one MMP family member activates another. ECM degradation and MMP activation can be initiated by several different stimuli including changes in ECM mechanical properties or intracellular contractility. These mechanical stimuli are known enhancers of metastatic potential. MMP-14 facilitates local ECM degradation and is well known as a major mediator of cell migration, angiogenesis and invasion. Recently, function blocking antibodies have been developed to specifically block MMP-14, providing a useful tool for research as well as therapeutic applications. Here we utilize a selective MMP-14 function blocking antibody to delineate the role of MMP-14 as an activator of other MMPs in response to changes in cellular contractility and ECM stiffness. Inhibition using function blocking antibodies reveals that MMP-14 activates soluble MMPs like MMP-2 and -9 under various mechanical stimuli in the pancreatic cancer cell line, Panc-1. In addition, inhibition of MMP-14 abates Panc-1 cell extension into 3D gels to levels seen with non-specific pan-MMP inhibitors at higher concentrations. This strengthens the case for MMP function blocking antibodies as more potent and specific MMP inhibition therapeutics.     

Time course of matrix metalloproteases and tissue inhibitors in bleomycin-induced pulmonary fibrosis

To investigate simultaneously localization and relative activity of MMPs during extracellular matrix (ECM) remodeling in bleomycin-induced pulmonary fibrosis in rat, we analyzed the time course of the expression, activity and/or concentration of gelatinases MMP-2 and MMP-9, collagenase MMP-1, matrylisin MMP-7, TIMP-1 and TIMP-2, both in alveolar space (cellular and extracellular compartments) and in lung tissue. MMP and TIMP expression was detected (immunohistochemistry) in lung tissue. MMP activity (zymography) and TIMP concentration (ELISA) were evaluated in lung tissue homogenate (LTH), BAL supernatant (BALs) and BAL cell pellet (BALp) 3, 7, 14, and 28 days after bleomycin intratracheal instillation. Immunohistochemistry showed an extensive MMP and TIMP expression from day 7 in a wide range of structural and inflammatory cells in treated rats. MMP-2 was present mainly in epithelia, MMP-9 in inflammatory cells. MMP-2 and MMP-9 activity was increased respectively in BAL fluid and BAL cells, with a peak at day 7. TIMP-1 and TIMP-2 concentration (ELISA) enhancement was delayed at day 14. In conclusion gelatinases and their inhibitors are significantly activated during bleomycin-induced pulmonary fibrosis. Marked changes in gelatinases activity are observed early in the alveolar compartment, with a prevailing extracellular activity of MMP-2 and a predominant intracellular distribution of MMP-9, while enzyme activity changes in lung parenchyma were less evident. In the repairing phase the reduction of gelatinases activity is synchronous with a peak of alveolar concentration of their inhibitors.     

PAI-1 in tissue fibrosis

Fibrosis is defined as a fibroproliferative or abnormal fibroblast activation-related disease. Deregulation of wound healing leads to hyperactivation of fibroblasts and excessive accumulation of extracellular matrix (ECM) proteins in the wound area, the pathological manifestation of fibrosis. The accumulation of excessive levels of collagen in the ECM depends on two factors: an increased rate of collagen synthesis and or decreased rate of collagen degradation by cellular proteolytic activities. The urokinase/tissue type plasminogen activator (uPA/tPA) and plasmin play significant roles in the cellular proteolytic degradation of ECM proteins and the maintenance of tissue homeostasis. The activities of uPA/tPA/plasmin and plasmin-dependent MMPs rely mostly on the activity of a potent inhibitor of uPA/tPA, plasminogen activator inhibitor-1 (PAI-1). Under normal physiologic conditions, PAI-1 controls the activities of uPA/tPA/plasmin/MMP proteolytic activities and thus maintains the tissue homeostasis. During wound healing, elevated levels of PAI-1 inhibit uPA/tPA/plasmin and plasmin-dependent MMP activities, and, thus, help expedite wound healing. In contrast to this scenario, under pathologic conditions, excessive PAI-1 contributes to excessive accumulation of collagen and other ECM protein in the wound area, and thus preserves scarring. While the level of PAI-1 is significantly elevated in fibrotic tissues, lack of PAI-1 protects different organs from fibrosis in response to injury-related profibrotic signals. Thus, PAI-1 is implicated in the pathology of fibrosis in different organs including the heart, lung, kidney, liver, and skin. Paradoxically, PAI-1 deficiency promotes spontaneous cardiac-selective fibrosis. In this review, we discuss the significance of PAI-1 in the pathogenesis of fibrosis in multiple organs.     

Release of Matrix Metalloproteinases-2 and 9 by S-Nitrosylated Caveolin-1 Contributes to Degradation of Extracellular Matrix in tPA-Treated Hypoxic Endothelial Cells

Intracranial hemorrhage remains the most feared complication in tissue plasminogen activator (tPA) thrombolysis for ischemic stroke. However, the underlying molecular mechanisms are still poorly elucidated. In this study, we reported an important role of caveolin-1 (Cav-1) s-nitrosylation in matrix metalloproteinase (MMP)-2 and 9 secretion from tPA-treated ischemic endothelial cells. Brain vascular endothelial cells (bEND3) were exposed to oxygen-glucose deprivation (OGD) for 2 h before adding recombinant human tPA for 6 h. This treatment induced a significant increase of MMP2 and 9 in the media of bEND3 cells and a simultaneous degradation of fibronectin and laminin β-1, the two main components of extracellular matrix (ECM). Inhibition of MMP2 and 9 with SB-3CT completely blocked the degradation of fibronectin and laminin β-1. ODG+tPA treatment led to Cav-1 shedding from bEND3 cells into the media. Notably, OGD triggered nitric oxide (NO) production and S-nitrosylationof Cav-1 (SNCav-1). Meanwhile tPA induced activation of ERK signal pathway and stimulates the secretion of SNCav-1. Pretreatment of bEND3 cells with C-PTIO (a NO scavenger) or U0126 (a specific ERK inhibitor) significantly reduced OGD-induced S-nitrosylation of Cav-1 in cells and blocked the secretion of Cav-1 and MMP2 and 9 into the media as well as the degradation of fibronectin and laminin β-1 in OGD and tPA-treated cells. These data indicate that OGD-triggered Cav-1 S-nitrosylation interacts with tPA-induced ERK activation to augment MMP2 and 9 secretion and subsequent ECM degradation, which may account for the exacerbation of ischemic blood brain barrier damage following tPA thrombolysis for ischemic stroke.     

Erythropoietin protects post-ischemic hearts by preventing extracellular matrix degradation: role of Jak2-ERK pathway

Factors predisposing to extracellular matrix degradation associated with myocardial ischemia/reperfusion (IR) usually cause cell death. Recombinant human erythropoietin (EPO) protects the myocardium from IR, but whether it affects extracellular matrix (ECM) degradation is not known. This study examined the effect of the Jak2-ERK pathway, which is triggered by EPO, on the expression of matrix metalloproteinases (MMPs), tissue inhibitor of MMP 4 (TIMP-4), and collagen in post-ischemic hearts. Rat hearts were isolated and perfused in a Langendorff apparatus. IR was induced by 40 min of stopped flow and 120 min of aerobic reperfusion; EPO was added immediately before reperfusion. Compared to untreated controls, poor recovery of the left ventricular developed pressure (LVDP) was seen in IR hearts. IR resulted in myocyte injury measured by creatine kinase MB release and infarction. Western blot analysis showed increased levels of MMP-2 and MMP-9 and reduced levels of TIMP-4 and collagen III. IR rats given 5 IU/ml of EPO showed improved LVDP with reduced injury. EPO increased Jak2 and ERK activity, decreased MMP expression, increased TIMP-4 expression, and prevented collagen degradation in IR hearts. All these effects were blocked by the upstream ERK inhibitor, U0126 (5 microM). These observations show that EPO attenuates extracellular matrix degradation following IR and this may be the basis of the protection from cell death. Jak2-ERK phosphorylation may be an important signal in this process.     

Spatial disruption and enhanced degradation of collagen with the transition from compensated ventricular hypertrophy to symptomatic congestive heart failure

The cardiac extracellular matrix (ECM) maintains the structural and mechanical integrity of the myocardium. We determined the alterations in the composition of the ECM coincident with the transition from compensated left ventricular (LV) hypertrophy (LVH) to symptomatic congestive heart failure (CHF) and the mechanisms underlying such changes. Heart failure was induced in ferrets by aortic banding. Myocardial collagen content was assessed by HPLC and histological analysis. Matrix metalloproteinase (MMP) activity and tissue inhibitor of metalloproteinase (TIMP) expression were evaluated using gelatin zymography and Western blotting, respectively. LV free wall thickness increased by 29% in asymptomatic LVH and was associated with a 20% increase in interstitial fibrosis (P < 0.05). CHF was coincident with increased plasma angiotensin II levels (149 +/- 48, 40 +/- 19, and 5.6 +/- 1 pg/ml for CHF, LVH, and sham, respectively; P < 0.01, CHF vs. sham and LVH), ventricular dilatation (LV internal diameter = 15 +/- 0.4 vs. 9 +/- 0.1 mm, P < 0.05), increased active MMP-9 (3.0- and 2.2-fold increase over sham and LVH, respectively, n = 5-10 animals per group, P < 0.01), and reduced myocardial total collagen content (3.5 +/- 0.4, 2.6 +/- 0.3, and 2.2 +/- 0.3% in sham, LVH, and CHF, respectively, P < 0.05). In CHF the distribution of collagen was markedly altered, becoming punctate in nature. No difference in MMP-2 activity, TIMP-1, TIMP-2, TIMP-3, or TIMP-4 expression, or collagen cross-linking was found at any time. The present work demonstrates structural reorganization and loss of collagen from cardiac ECM during the transition to decompensated CHF. The enhanced MMP-9 activity coincident with the transition to CHF provides potential therapeutic opportunities for managing the progression from asymptomatic LVH to symptomatic CHF.     

Matrix metalloproteinase expression and activity following prostaglandin F(2 alpha)-induced luteolysis

Luteal tissue contains matrix metalloproteinases (MMPs) that cleave specific components of the extracellular matrix (ECM) and are inhibited by tissue inhibitors of metalloproteinases (TIMPs). We previously reported a decrease in luteal TIMP-1 within 15 min of prostaglandin F(2 alpha) (PGF(2 alpha))-induced luteolysis. An increase in the MMP:TIMP ratio may promote ECM degradation and apoptosis, as observed in other tissues that undergo involution. The objectives of these experiments were to determine whether 1) PGF(2 alpha) affects expression of mRNA encoding fibrillar collagenases (MMP-1 and -13), gelatinases A and B (MMP-2 and -9), membrane type (mt)-1 MMP (MMP-14), stromelysin (MMP-3), and matrilysin (MMP-7), and 2) PGF(2 alpha) increases MMP activity during PGF(2 alpha)-induced luteolysis in sheep. Corpora lutea (n = 3-10/time point) were collected at 0, 15, and 30 min and 1, 2, 4, 6, 12, 24, and 48 h after PGF(2 alpha) administration. Northern blot analysis confirmed the presence of all MMPs except MMP-9. Expression of mRNA for the above MMPs (except MMP-2) increased significantly (P < 0.05) by 30 min, and all MMPs increased significantly (P < 0.05) by 6 h after PGF(2 alpha) administration. Expression of MMP-14 mRNA increased significantly (P < 0.05) by 15 min post-PGF(2 alpha) and remained elevated through 48 h. MMP activity in luteal homogenates (following proenzyme activation and inactivation of inhibitors) was increased significantly (P < 0.05) by 15 min and remained elevated through 48 h post-PGF(2 alpha). MMP activity was localized (in situ zymography) to the pericellular area of various cell types in the 0-h group and was markedly increased by 30 min post-PGF(2 alpha). MMP mRNA expression and activity were significantly increased following PGF(2 alpha) treatment. Increased MMP activity may promote ECM degradation during luteolysis.     

MMP Secretion Rate and Inter-invadopodia Spacing Collectively Govern Cancer Invasiveness

Invadopodia are micron-sized invasive structures that mediate extracellular matrix (ECM) degradation through a combination of membrane-bound and soluble matrix metalloproteinases (MMPs). However, how such localized degradation is converted into pores big enough for cancer cells to invade, and the relative contributions of membrane-bound versus soluble MMPs to this process remain unclear. In this article, we address these questions by combining experiments and simulations. We show that in MDA-MB-231 cells, an increase in ECM density enhances invadopodia-mediated ECM degradation and decreases inter-invadopodia spacing. ECM degradation is mostly mediated by soluble MMPs, which are activated by membrane-bound MT1-MMP. We present a computational model of invadopodia-mediated ECM degradation, which recapitulates the above observations and identifies MMP secretion rate as an important regulator of invadopodia stability. Simulations with multiple invadopodia suggest that inter-invadopodia spacing and MMP secretion rate collectively dictate the size of the degraded zones. Taken together, our results suggest that for creating pores conducive for cancer invasion, cells must tune inter-invadopodia spacing and MMP secretion rate in an ECM density-dependent manner, thereby striking a balance between invadopodia penetration and ECM degradation.     

Mechanical stress regulates the mechanotransduction and metabolism of cardiac fibroblasts in fibrotic cardiac diseases

Fibrotic cardiac diseases are characterized by myocardial fibrosis that results in maladaptive cardiac remodeling. Cardiac fibroblasts (CFs) are the main cell type responsible for fibrosis. In response to stress or injury, intrinsic CFs develop into myofibroblasts and produce excess extracellular matrix (ECM) proteins. Myofibroblasts are mechanosensitive cells that can detect changes in tissue stiffness and respond accordingly. Previous studies have revealed that some mechanical stimuli control fibroblast behaviors, including ECM formation, cell migration, and other phenotypic traits. Further, metabolic alteration is reported to regulate fibrotic signaling cascades, such as the transforming growth factor-β pathway and ECM deposition. However, the relationship between metabolic changes and mechanical stress during fibroblast-to-myofibroblast transition remains unclear. This review aims to elaborate on the crosstalk between mechanical stress and metabolic changes during the pathological transition of cardiac fibroblasts.     

Impact of 4‐hydroxynonenal on matrix metalloproteinase‐9 regulation in lipopolysaccharide‐stimulated RAW 264.7 cells

Tissue degradation and leukocyte extravasation suggest proteolytic destruction of the extracellular matrix (ECM) during severe malaria. Matrix metalloproteinases (MMPs) play an established role in ECM turnover, and increased MMP‐9 protein abundance is correlated with malarial infection. The malaria pigment hemozoin (Hz) is a heme detoxification biomineral that is produced during infection and associated with biologically active lipid peroxidation products such as 4‐hydroxynonenal (HNE) adsorbed to its surface. Hz has innate immunomodulatory activity, and many of its effects can be reproduced by exogenously added HNE. Hz phagocytosis enhances MMP‐9 expression in monocytes; thus, this study was designed to examine the ability of HNE to alter MMP‐9 regulation in activated cells of macrophage lineage. Data show that treatment of lipopolysaccharide‐stimulated RAW 264.7 cells with HNE increased MMP‐9 secretion and activity. HNE treatment abolished the cognate tissue inhibitor of metalloproteinase‐1 protein levels, further decreasing MMP‐9 regulation. Phosphorylation of both p38 mitogen‐activated protein kinase (MAPK) and c‐Jun NH2‐terminal kinase was induced by HNE, but only p38 MAPK inhibition lessened MMP‐9 secretion. These results demonstrate the in vitro ability of HNE to cause MMP‐9 dysregulation in an activated cell model. The findings may extend to myriad pathologies associated with lipid peroxidation and elevated MMP‐9 levels leading to tissue damage. Copyright © 2015 John Wiley & Sons, Ltd.