Biochimica et Biophysica Acta (BBA) - General Subjects
ReviewThrombospondin-2 and extracellular matrix assembly☆
Introduction
The extracellular matrix (ECM) consists of numerous proteins, including several collagen types, fibronectin, and laminin, which are arranged into a 3D network. Within the network structure of the ECM, other components such as proteoglycans (PG) contribute to its integrity and organization [1]. Alteration of major ECM proteins due to mutation, lack of translation, improper folding, or improper assembly, leads to significant changes in ECM properties. Clinically, Marfan's syndrome, Ehlers–Danlos syndrome, and Stickler syndrome result from mutations in one or more of the collagen genes or collagen-processing enzymes [2]. Similarly, non-structural ECM components such as matricellular proteins play a major role in influencing ECM assembly. Matricellular proteins do not contribute directly to the stable structure of the ECM; however, they possess cellular binding sites that influence cell–matrix interactions and cellular signaling/behavior [3]. In addition, they are also able to sequester growth factors and modulate their bioavailability [4].
Even though the assembly of ECM proteins into a cohesive, functional matrix is integral for vertebral life, it is not fully understood how all components come together in an organized network. However, for specific matrix components, much has been determined regarding the transition from monomeric subunits to individual multimeric proteins. For example, collagen I (and other fibrillar collagens) has a multi-step self-assembly process that includes side by side alignment of triple helical tropocollagen monomers, the formation of microfibrillar units, the fusion of these microfibrillar units into larger fibrils, and continued fusion that leads to eventual formation of micrometer sized fibers that display collagen's characteristic banding pattern and periodicity [5]. Proteoglycans, glycosaminoglycans, and minor collagen types also participate in fibrillogenesis. Fibronectin also displays a unique, step-wise, self-assembly process that is characterized by integrin-dependence and necessary actin cytoskeletal connections [6]. In addition to intramolecular interactions, ECM proteins utilize specific recognition sequences to bind to each other and contribute to matrix formation and stabilization. For example, fibronectin and SPARC bind collagen, and thrombospondin (TSP) 1 can bind laminin. Moreover, post-translational modifications such as cross-linking, glycosylation, and hydroxylation have been shown to influence ECM properties [1], [7], [8].
The TSPs are a family of five matricellular proteins, divided into groups based on their trimeric (TSP1 and TSP2) or pentameric (TSP3, TSP4, and TSP5) structure [9]. TSP1 and TSP2 are of particular interest in wound repair and tissue remodeling due to their anti-angiogenic activity. TSP1 is produced primarily by platelets and has been shown to be integral in the acute inflammatory phase of tissue repair [10]. Unlike TSP1, TSP2 is primarily produced by fibroblasts and smooth muscle cells, and is implicated in the later, remodeling phases of tissue repair [11]. TSP1 and TSP2 present 85% amino acid and structural conservation, thus, TSP2 has similar binding domains to TSP1 [12]. Both proteins are synthesized and secreted as homotrimers consisting of N-terminal laminin G-like domain, a vWF pro-collagen-like domain, three type 1 properdin-like repeats, three EGF-like type II repeats, seven type III repeats, and a carboxy terminal lectin type domain (Fig. 1). These domains are involved in multiple interactions with cell surface receptors (LRP, CD36, CD 47, and numerous integrins), ECM components (decorin, fibronectin, HSPGs), growth factors (TGF-b, FGF2), enzymes (MMPs, elastase, cathepsin G), and calcium binding. The relative significance of specific TSP1/2 domain interactions in various cell functions and the molecular basis for the regulation of angiogenesis have been reviewed recently [13], [14]. In this review, we will explore the role of TSP2 as a modulator of cell–matrix interactions and ECM assembly. In addition, we will focus on the novel properties of the TSP2-null ECM and its potential for therapeutic applications.
Section snippets
Connective tissue abnormalities
TSP2-null mice, generated over a decade ago, displayed normal physical appearance and reproduced at the expected Mendelian ratio. However, upon handling, it was noticed that the mice had lax skin and connective tissues including ligaments and tendons [15]. Tensile strength analysis of skin showed increased fragility and laxity. Further examination revealed a bleeding diathesis, increase in bone density, and increase in small to medium sized blood vessels. Interestingly, it was determined via
Characterization and properties of TSP2-null ECM
As mentioned above, it was noted that TSP2-null mice have lax tendons and ligaments and increased skin fragility suggestive of abnormal collagen fibrillogenesis. Electron microscopy analysis of the skin and tail tendons from TSP2-null mice revealed disorganized collagen fiber weave and collagen fibrils with uneven contours and larger diameters compared to that of WT mice [15]. These findings highlighted that TSP2 is necessary for the proper formation and organization of both collagen fibrils
TSP2 and cardiac ECM integrity
Is matrix-incorporated TSP2 always a villain, however? What about instances when matrix structural integrity and architecture need to be maintained? In myocardial remodeling, such as during left ventricular hypertrophy and the progression of cardiomyopathy, both the cardiac cells and ECM experience large amounts of stress. Surprisingly, TSP2 has been shown to be cardioprotective in many instances. Scroen et al. demonstrated that while chronically elevated levels of TSP2 are detrimental and
Mouse models with abnormal collagen/ECM phenotypes
Other prominent ECM components such as decorin, lumican, and fibromodulin have been shown to produce similar phenotypes when knocked-out in murine models. Decorin, a member of the small leucine-rich proteoglycan (SLRP) family and aptly named for its “decoration” of collagen fibrils, has been demonstrated to influence fibrillogenesis, matrix assembly, and growth factor availability [50]. It has also been shown that decorin, in soluble form, acts as a signaling proteoglycan through tyrosine
Cell-derived ECM as a potential substrate for in vitro studies and in vivo applications
Decellularization of tissues and cell cultures has allowed for tissue- and cell-derived ECM to be employed as a bioactive substrate to investigate cell function [64], [65]. Methods of decellularization can be broken down into three main categories: chemical, enzymatic, and physical, with combinations of these methods often used in various protocols. Chemical decellularization methods tend to employ a mixture of basic and hypo/hypertonic solutions, plus ionic, non-ionic, or zwitterionic
Acknowledgements
This work was supported by NIH (HL107205) and the Gruber Science Fellowship (to N.E.C).
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This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties.