Oxidative damage to extracellular matrix and its role in human pathologies

The extracellular compartments of most biological tissues are significantly less well protected against oxidative damage than intracellular sites and there is considerable evidence for such compartments being subject to a greater oxidative stress and an altered redox balance. However, with some notable exceptions (e.g., plasma and lung lining fluid) oxidative damage within these compartments has been relatively neglected and is poorly understood. In particular information on the nature and consequences of damage to extracellular matrix is lacking despite the growing realization that changes in matrix structure can play a key role in the regulation of cellular adhesion, proliferation, migration, and cell signaling. Furthermore, the extracellular matrix is widely recognized as being a key site of cytokine and growth factor binding, and modification of matrix structure might be expected to alter such behavior. In this paper we review the potential sources of oxidative matrix damage, the changes that occur in matrix structure, and how this may affect cellular behavior. The role of such damage in the development and progression of inflammatory diseases is discussed.     

Oxidation of glycosaminoglycans by free radicals and reactive oxidative species: A review of investigative methods

Abstract

Glycosaminoglycans, in particular hyaluronan (HA), and proteoglycans are components of the extracellular matrix (ECM). The ECM plays a key role in the regulation of cellular behaviour and alterations to it can modulate both the development of human diseases as well as controlling normal biochemical processes such as cell signalling and pro-inflammatory responses. For these reasons, in vitro fragmentation studies of glycosaminoglycans by free radicals and oxidative species are seen to be relevant to the understanding of in vivo studies of damage to the ECM. A wide range of investigative techniques have therefore been applied to gain insights into the relative fragmentation effects of several reactive oxidative species with the ultimate goal of determining mechanisms of fragmentation at the molecular level. These methods are reviewed here.     

促进老年退行性骨关节炎软骨内源性修复的化合物研究进展

老年退行性骨关节炎(OA)是由关节损伤、肥胖和衰老等因素引起的一种退行性疾病,最终引起关节软骨损伤,导致运动功能障碍。软骨细胞及细胞外基质是软骨组织的主要成分,它们的损伤是引起OA的根本原因。目前OA的治疗仅限于缓解症状,而随着干细胞的发现及对软骨细胞的深入认识,开发增强软骨内源性修复的药物是OA治疗的重要方向。目前研究发现,kartogenin等化合物可以促进间充质干细胞选择性的分化为软骨细胞而起到修复作用,此外,一些化合物还可以调控软骨细胞的信号通路,起到促进软骨细胞增殖、抑制软骨细胞凋亡、抑制基质金属蛋白酶活性、增加细胞外基质合成等作用,从而维持软骨细胞的数量、促进软骨基质的合成而抑制其降解。这些方法比常规通过微创刺激内源性干细胞或移植自体细胞更加安全、有效。本文就化合物对促进老年退行性骨炎软骨内源性修复的研究进行综述,为发现更多的有效化合物提供基础。     

Impact of Mycobacterium ulcerans biofilm on transmissibility to ecological niches and Buruli ulcer pathogenesis

The role of biofilms in the pathogenesis of mycobacterial diseases remains largely unknown. Mycobacterium ulcerans, the etiological agent of Buruli ulcer, a disfiguring disease in humans, adopts a biofilm-like structure in vitro and in vivo, displaying an abundant extracellular matrix (ECM) that harbors vesicles. The composition and structure of the ECM differs from that of the classical matrix found in other bacterial biofilms. More than 80 proteins are present within this extracellular compartment and appear to be involved in stress responses, respiration, and intermediary metabolism. In addition to a large amount of carbohydrates and lipids, ECM is the reservoir of the polyketide toxin mycolactone, the sole virulence factor of M. ulcerans identified to date, and purified vesicles extracted from ECM are highly cytotoxic. ECM confers to the mycobacterium increased resistance to antimicrobial agents, and enhances colonization of insect vectors and mammalian hosts. The results of this study support a model whereby biofilm changes confer selective advantages to M. ulcerans in colonizing various ecological niches successfully, with repercussions for Buruli ulcer pathogenesis.     

Distribution of extracellular matrix proteins type I collagen, type IV collagen, fibronectin, and laminin in mouse folliculogenesis

The extracellular matrix (ECM) plays a prominent role in ovarian function by participating in processes such as cell migration, proliferation, growth, and development. Although some of these signaling processes have been characterized in the mouse, the relative quantity and distribution of ECM proteins within developing follicles of the ovary have not been characterized. This study uses immunohistochemistry and real-time PCR to characterize the ECM components type I collagen, type IV collagen, fibronectin, and laminin in the mouse ovary according to follicle stage and cellular compartment. Collagen I was present throughout the ovary, with higher concentrations in the ovarian surface epithelium and follicular compartments. Collagen IV was abundant in the theca cell compartment with low-level expression in the stroma and granulosa cells. The distribution of collagen was consistent throughout follicle maturation. Fibronectin staining in the stroma and theca cell compartment increased throughout follicle development, while staining in the granulosa cell compartment decreased. Heavy staining was also observed in the follicular fluid of antral follicles. Laminin was localized primarily to the theca cell compartment, with a defined ring at the exterior of the follicular granulosa cells marking the basement membrane. Low levels of laminin were also apparent in the stroma and granulosa cell compartment. Taken together, the ECM content of the mouse ovary changes during follicular development and reveals a distinct spatial and temporal pattern. This understanding of ECM composition and distribution can be used in the basic studies of ECM function during follicle development, and could aid in the development of in vitro systems for follicle growth.     

Integrin–ECM interactions and membrane-associated Catalase cooperate to promote resilience of the Drosophila intestinal epithelium

Balancing cellular demise and survival constitutes a key feature of resilience mechanisms that underlie the control of epithelial tissue damage. These resilience mechanisms often limit the burden of adaptive cellular stress responses to internal or external threats. We recently identified Diedel, a secreted protein/cytokine, as a potent antagonist of apoptosis-induced regulated cell death in the Drosophila intestinal midgut epithelium during aging. Here, we show that Diedel is a ligand for RGD-binding Integrins and is thus required for maintaining midgut epithelial cell attachment to the extracellular matrix (ECM)-derived basement membrane. Exploiting this function of Diedel, we uncovered a resilience mechanism of epithelial tissues, mediated by Integrin–ECM interactions, which shapes cell death spreading through the regulation of cell detachment and thus cell survival. Moreover, we found that resilient epithelial cells, enriched for Diedel–Integrin–ECM interactions, are characterized by membrane association of Catalase, thus preserving extracellular reactive oxygen species (ROS) balance to maintain epithelial integrity. Intracellular Catalase can relocalize to the extracellular membrane to limit cell death spreading and repair Integrin–ECM interactions induced by the amplification of extracellular ROS, which is a critical adaptive stress response. Membrane-associated Catalase, synergized with Integrin–ECM interactions, likely constitutes a resilience mechanism that helps balance cellular demise and survival within epithelial tissues.

A key feature of the resilience mechanisms that underlie the control of epithelial tissue damage is the balance between cell death and survival. This study shows that the anti-oxidant enzyme catalase can relocate to membranes in order to promote the resilience of the Drosophila midgut epithelium, synergizing with integrin-ECM interactions to prevent the spread of cell death.     

Mesenchymal stem cell-conditioned media recovers lung fibroblasts from cigarette smoke-induced damage

Cigarette smoking causes apoptotic death, senescence, and impairment of repair functions in lung fibroblasts, which maintain the integrity of alveolar structure by producing extracellular matrix (ECM) proteins. Therefore, recovery of lung fibroblasts from cigarette smoke-induced damage may be crucial in regeneration of emphysematous lung resulting from degradation of ECM proteins and subsequent loss of alveolar cells. Recently, we reported that bone marrow-derived mesenchymal stem cell-conditioned media (MSC-CM) led to angiogenesis and regeneration of lung damaged by cigarette smoke. In this study, to further investigate reparative mechanisms for MSC-CM-mediated lung repair, we attempted to determine whether MSC-CM can recover lung fibroblasts from cigarette smoke-induced damage. In lung fibroblasts exposed to cigarette smoke extract (CSE), MSC-CM, not only inhibited apoptotic death, but also induced cell proliferation and reversed CSE-induced changes in the levels of caspase-3, p53, p21, p27, Akt, and p-Akt. MSC-CM also restored expression of ECM proteins and collagen gel contraction while suppressing CSE-induced expression of cyclooxygenase-2 and microsomal PGE(2) synthase-2. The CSE-opposing effects of MSC-CM on cell fate, expression of ECM proteins, and collagen gel contraction were partially inhibited by LY294002, a phosphatidylinositol 3-kinase (PI3K) inhibitor. In rats, MSC-CM administration also resulted in elevation of p-Akt and restored proliferation of lung fibroblasts, which was suppressed by exposure to cigarette smoke. Taken together, these data suggest that MSC-CM may recover lung fibroblasts from cigarette smoke-induced damage, possibly through inhibition of apoptosis, induction of proliferation, and restoration of lung fibroblast repair function, which are mediated in part by the PI3K/Akt pathway.     

Localizing matrix metalloproteinase activities in the pericellular environment

Matrix metalloproteinases (MMPs) are a group of structurally related proteolytic enzymes containing a zinc ion in the active site. They are secreted from cells or bound to the plasma membrane and hydrolyze extracellular matrix (ECM) and cell surface-bound molecules. They therefore play key roles in morphogenesis, wound healing, tissue repair and remodeling in diseases such as cancer and arthritis. Although the cell anchored membrane-type MMPs (MT-MMPs) function pericellularly, the secreted MMPs have been considered to act within the ECM, away from the cells from which they are synthesized. However, recent studies have shown that secreted MMPs bind to specific cell surface receptors, membrane-anchored proteins or cell-associated ECM molecules and function pericellularly at focussed locations. This minireview describes examples of cell surface and pericellular partners of MMPs, as well as how they alter enzyme function and cellular behaviour.     

Free radical studies of components of the extracellular matrix: contributions to protection of biomolecules and biomaterials from sterilising doses of ionising radiation

The purpose of the current review is show how the principles and techniques of radiation chemistry have enabled the direct reactions of free radicals with biomolecules and biomaterials to be investigated at the molecular level. In particular, the review focusses on the free radical-induced fragmentation of glycosaminoglycans. Glycosaminoglycans are large linear polysaccharides consisting of repeating disaccharide units and are important components of the extracellular matrix (ECM) either in free form (hyaluronan) or as a component of proteoglycans. Oxidative damage of the extracellular matrix components by either enzymatic or non-enzymatic pathways may have implications for the initiation and progression of a range of human diseases. These include arthritis, kidney disease, cardiovascular disease, lung disease, periodontal disease and chronic inflammation. Oxidative damage to hyaluronan by reactive oxidative species and thus the potential mechanism of damage to the ECM and its role in human pathologies is reviewed with particular focus on damage initiated by potential in vivo free radicals such as superoxide, carbonate and hydroxyl radicals. Such knowledge has also allowed radiation protecting systems to be developed so that sterilising doses of radiation can be delivered to sensitive biomolecules such as proteins and glycosaminoglycans, and also to sensitive biomaterials such as tissue allografts.     

Biological and metabolic response in STS-135 space-flown mouse skin

Abstract

There is evidence that space flight condition-induced biological damage is associated with increased oxidative stress and extracellular matrix (ECM) remodeling. To explore possible mechanisms, changes in gene expression profiles implicated in oxidative stress and in ECM remodeling in mouse skin were examined after space flight. The metabolic effects of space flight in skin tissues were also characterized. Space Shuttle Atlantis (STS-135) was launched at the Kennedy Space Center on a 13-day mission. Female C57BL/6 mice were flown in the STS-135 using animal enclosure modules (AEMs). Within 3–5 h after landing, the mice were euthanized and skin samples were harvested for gene array analysis and metabolic biochemical assays. Many genes responsible for regulating production and metabolism of reactive oxygen species (ROS) were significantly (p < 0.05) altered in the flight group, with fold changes >1.5 compared to AEM control. For ECM profile, several genes encoding matrix and metalloproteinases involved in ECM remodeling were significantly up-/down-regulated following space flight. To characterize the metabolic effects of space flight, global biochemical profiles were evaluated. Of 332 named biochemicals, 19 differed significantly (p < 0.05) between space flight skin samples and AEM ground controls, with 12 up-regulated and 7 down-regulated including altered amino acid, carbohydrate metabolism, cell signaling, and transmethylation pathways. Collectively, the data demonstrated that space flight condition leads to a shift in biological and metabolic homeostasis as the consequence of increased regulation in cellular antioxidants, ROS production, and tissue remodeling. This indicates that astronauts may be at increased risk for pathophysiologic damage or carcinogenesis in cutaneous tissue.     

Fibroblast adhesion results in the induction of a matrix remodeling gene expression program

Fibrosis is believed to occur through the failure to terminate the normal tissue remodeling program. Tissue repair intimately involves the ability of fibroblasts to attach to extracellular matrix (ECM), resulting in cell migration and ECM contraction. Elevated, activated adhesive signaling is a key phenotypic hallmark of fibrotic cells. The precise contribution of adhesion to tissue remodeling and repair and fibrotic responses in fibroblasts is unclear, but involves focal adhesion kinase (FAK). FAK signals downstream of integrin-mediates attachment of fibroblasts to extracellular matrix. In this report, we show that FAK is required for the expression of a cohort of mRNAs encoding ECM and matrix remodeling genes including CCN2, alpha-smooth muscle actin (SMA) and type I collagen. Adhesion of fibroblasts to fibronectin, a component of the provisional matrix deposited in the initial phases of tissue repair, also resulted in the induction of CCN2, alpha-SMA and type I collagen mRNAs. Endothelin-1 (ET-1), a key inducer of pro-fibrotic gene expression, was also induced upon fibroblast attachment to ECM, and antagonism of the ET-1 receptors significantly reduced the ability of adhesion to induce expression of CCN2, alpha-SMA and type I collagen mRNAs. These results suggest that adhesion of fibroblasts to matrix during the initial phases of tissue remodeling and repair may actively contribute to the tissue repair program through the induction of pro-fibrotic gene expression.     

Materials as morphogenetic guides in tissue engineering

Within native tissues cells are held within the extracellular matrix (ECM), which has a role in maintaining homeostasis, guiding development and directing regeneration. Efforts in tissue engineering have aimed to mimick the ECM to help guide morphogenesis and tissue repair. Studies have not only looked at ways to mimick the structure and characteristics of the ECM, but have also considered ways to reproduce its molecular properties including its bioadhesive character, proteolytic susceptibility and ability to bind growth factors.     

Extracellular matrix (ECM) microstructural composition regulates local cell-ECM biomechanics and fundamental fibroblast behavior: a multidimensional perspective.

The extracellular matrix (ECM) provides the principal means by which mechanical information is communicated between tissue and cellular levels of function. These mechanical signals play a central role in controlling cell fate and establishing tissue structure and function. However, little is known regarding the mechanisms by which specific structural and mechanical properties of the ECM influence its interaction with cells, especially within a tissuelike context. This lack of knowledge precludes formulation of biomimetic microenvironments for effective tissue repair and replacement. The present study determined the role of collagen fibril density in regulating local cell-ECM biomechanics and fundamental fibroblast behavior. The model system consisted of fibroblasts seeded within collagen ECMs with controlled microstructure. Confocal microscopy was used to collect multidimensional images of both ECM microstructure and specific cellular characteristics. From these images temporal changes in three-dimensional cell morphology, time- and space-dependent changes in the three-dimensional local strain state of a cell and its ECM, and spatial distribution of beta1-integrin were quantified. Results showed that fibroblasts grown within high-fibril-density ECMs had decreased length-to-height ratios, increased surface areas, and a greater number of projections. Furthermore, fibroblasts within low-fibril-density ECMs reorganized their ECM to a greater extent, and it appeared that beta1-integrin localization was related to local strain and ECM remodeling events. Finally, fibroblast proliferation was enhanced in low-fibril-density ECMs. Collectively, these results are significant because they provide new insight into how specific physical properties of a cell's ECM microenvironment contribute to tissue remodeling events in vivo and to the design and engineering of functional tissue replacements.     

Degradation of extracellular matrix by peroxynitrite/peroxynitrous acid

The extracellular matrix (ECM) provides strength and elasticity to tissues and plays a key role in regulating cell behavior; damage to this material is believed to be a major factor in many inflammatory diseases. Peroxynitrite/peroxynitrous acid, which is generated at elevated levels at sites of inflammation, is believed to play a role in ECM damage; however, the mechanisms involved are poorly understood. Here we examined the reactions of bolus peroxynitrite, and that generated in a time-dependent manner by SIN-1 decomposition, with ECM isolated from a vascular smooth muscle cell line and porcine thoracic aorta. Bolus peroxynitrite caused the release of ECM glycosaminoglycans and proteins, the formation of 3-nitroTyr, and the detection of ECM-derived radicals (by immuno-spin trapping) in a concentration-dependent manner. Release and nitration of ECM components were modulated by the local pH and bicarbonate. SIN-1 caused the release of glycosaminoglycan, but not protein, from vascular smooth muscle cell-derived ECM in a concentration-, time-, and pH-dependent manner. The data presented here suggest that peroxynitrite-mediated damage to ECM occurs via a radical-mediated pathway. These reactions may contribute to ECM damage at sites of inflammation and play a role in disease progression, including rupture of atherosclerotic lesions.     

Smad3 null mice develop airspace enlargement and are resistant to TGF-beta-mediated pulmonary fibrosis

Transforming growth factor-beta 1 plays a key role in the pathogenesis of pulmonary fibrosis, mediating extracellular matrix (ECM) gene expression through a series of intracellular signaling molecules, including Smad2 and Smad3. We show that Smad3 null mice (knockout (KO)) develop progressive age-related increases in the size of alveolar spaces, associated with high spontaneous presence of matrix metalloproteinases (MMP-9 and MMP-12) in the lung. Moreover, transient overexpression of active TGF-beta 1 in lungs, using adenoviral vector-mediated gene transfer, resulted in progressive pulmonary fibrosis in wild-type mice, whereas no fibrosis was seen in the lungs of Smad3 KO mice up to 28 days. Significantly higher levels of matrix components (procollagen 3A1, connective tissue growth factor) and antiproteinases (plasminogen activator inhibitor-1, tissue inhibitor of metalloproteinase-1) were detected in wild-type lungs 4 days after TGF-beta 1 administration, while no such changes were seen in KO lungs. These data suggest a pivotal role of the Smad3 pathway in ECM metabolism. Basal activity of the pathway is required to maintain alveolar integrity and ECM homeostasis, but excessive signaling through the pathway results in fibrosis characterized by inhibited degradation and enhanced ECM deposition. The Smad3 pathway is involved in pathogenic mechanisms mediating tissue destruction (lack of repair) and fibrogenesis (excessive repair).     

A bit of give and take: the relationship between the extracellular matrix and the developing chondrocyte

The extracellular matrix (ECM), once thought to be a static structural component of tissues, is now known to play a complex and dynamic role in a variety of cellular functions in a number of diverse tissues. A significant body of literature attests to the ability of the ECM to communicate both spatial and temporal information to adherent cells, thereby directing cell behavior via interactions between the ECM and cell-surface receptors. Moreover, volumes of experimental data show that a great deal of communication travels in the opposite direction, from the cell to the ECM, allowing for regulation of the cues transmitted by the ECM. As such, the ECM, with respect to its components and their organization, is not a fixed reflection of the state the local microenvironment in which a cell finds itself at a particular time, but rather is able to respond to and effect changes in its local microenvironment. As an example of the developmental consequences of ECM interactions, this review gives an overview of the 'give and take' relationship between the ECM and the cells of the developing skeletal elements, in particular, the chondrocyte.     

Human disease-associated extracellular matrix orthologs ECM3 and QBRICK regulate primary mesenchymal cell migration in sea urchin embryos

Sea urchin embryos have been one of model organisms to investigate cellular behaviors because of their simple cell composition and transparent body. They also give us an opportunity to investigate molecular functions of human proteins of interest that are conserved in sea urchin. Here we report that human disease-associated extracellular matrix orthologues ECM3 and QBRICK are necessary for mesenchymal cell migration during sea urchin embryogenesis. Immunofluorescence has visualized the colocalization of QBRICK and ECM3 on both apical and basal surface of ectoderm. On the basal surface, QBRICK and ECM3 constitute together a mesh-like fibrillar structure along the blastocoel wall. When the expression of ECM3 was knocked down by antisense-morpholino oligonucleotides, the ECM3-QBRICK fibrillar structure completely disappeared. When QBRICK was knocked down, the ECM3 was still present, but the basally localized fibers became fragmented. The ingression and migration of primary mesenchymal cells were not critically affected, but their migration at later stages was severely affected in both knock-down embryos. As a consequence of impaired primary mesenchymal cell migration, improper spicule formation was observed. These results indicate that ECM3 and QBRICK are components of extracellular matrix, which play important role in primary mesenchymal cell migration, and that sea urchin is a useful experimental animal model to investigate human disease-associated extracellular matrix proteins.