SPARC, a matricellular protein: at the crossroads of cell-matrix communication.

SPARC is a multifunctional glycoprotein that belongs to the matricellular group of proteins. It modulates cellular interaction with the extracellular matrix (ECM) by its binding to structural matrix proteins, such as collagen and vitronectin, and by its abrogation of focal adhesions, features contributing to a counteradhesive effect on cells. SPARC inhibits cellular proliferation by an arrest of cells in the G1 phase of the cell cycle. It also regulates the activity of growth factors, such as platelet-derived growth factor (PDGF), fibroblast growth factor (FGF)-2, and vascular endothelial growth factor (VEGF). The expression of SPARC in adult animals is limited largely to remodeling tissue, such as bone, gut mucosa, and healing wounds, and it is prominent in tumors and in disorders associated with fibrosis. The crystal structure of two of the three domains of the protein has revealed a novel follistatin-like module and an extracellular calcium-binding (EC) module containing two EF-hand motifs. The follistatin-like module and the EC module are shared by at least four other proteins that comprise a family of SPARC-related genes. Targeted disruption of the SPARC locus in mice has shown that SPARC is important for lens transparency, as SPARC-null mice develop cataracts shortly after birth. SPARC is a prototypical matricellular protein that functions to regulate cell-matrix interactions and thereby influences many important physiological and pathological processes.     

SPARC, a matricellular protein: at the crossroads of cell–matrix communication: [Matrix Biology (2000) 569–580]

The publisher regrets that the above article was published with several typographical errors. The corrected version appears on the following pages. SPARC is a multifunctional glycoprotein that belongs to the matricellular group of proteins. It modulates cellular interaction with the extracellular matrix (ECM) by its binding to structural matrix proteins, such as collagen and vitronectin, and by its abrogation of focal adhesions, features contributing to a counteradhesive effect on cells. SPARC inhibits cellular proliferation by an arrest of cells in the G1 phase of the cell cycle. It also regulates the activity of growth factors, such as platelet-derived growth factor (PDGF), fibroblast growth factor (FGF)-2, and vascular endothelial growth factor (VEGF). The expression of SPARC in adult animals is limited largely to remodeling tissue, such as bone, gut mucosa, and healing wounds, and it is prominent in tumors and in disorders associated with fibrosis. The crystal structure of two of the three domains of the protein has revealed a novel follistatin-like module and an extracellular calcium-binding (EC) module containing two EF-hand motifs. The follistatin-like module and the EC module are shared by at least four other proteins that comprise a family of SPARC-related genes. Targeted disruption of the SPARC locus in mice has shown that SPARC is important for lens transparency, as SPARC-null mice develop cataracts shortly after birth. SPARC is a prototypical matricellular protein that functions to regulate cell–matrix interactions and thereby influences many important physiological and pathological processes.     

SPARC, a matricellular glycoprotein with important biological functions.

SPARC (secreted protein, acidic and rich in cysteine) is a unique matricellular glycoprotein that is expressed by many different types of cells and is associated with development, remodeling, cell turnover, and tissue repair. Its principal functions in vitro are counteradhesion and antiproliferation, which proceed via different signaling pathways. SPARC consists of three domains, each of which has independent activity and unique properties. The extracellular calcium binding module and the follistatin-like module have been recently crystallized. Specific interactions between SPARC and growth factors, extracellular matrix proteins, and cell surface proteins contribute to the diverse activities described for SPARC in vivo and in vitro. The location of SPARC in the nuclear matrix of certain proliferating cells, but only in the cytosol of postmitotic neurons, indicates potential functions of SPARC as a nuclear protein, which might be involved in the regulation of cell cycle progression and mitosis. High levels of SPARC have been found in adult eye, and SPARC-null mice exhibit cataracts at 1-2 months of age. This animal model provides an excellent opportunity to confirm and explore some of the properties of SPARC, to investigate cataractogenesis, and to study SPARC-related family proteins, e.g., SC1/hevin, a counteradhesive matricellular protein that might functionally compensate for SPARC in certain tissues.(J Histochem Cytochem 47:1495-1505, 1999)     

SPARC and tumor growth: Where the seed meets the soil?

Matricellular proteins mediate interactions between cells and their extracellular environment. This functional protein family includes several structurally unrelated members, such as SPARC, thrombospondin 1, tenascin C, and osteopontin, as well as some homologs of these proteins, such as thrombospondin 2 and tensascin X. SPARC, a prototypic matricellular protein, and its homolog hevin, have deadhesive effects on cultured cells and have been characterized as antiproliferative factors in some cellular contexts. Both proteins are produced at high levels in many types of cancers, especially by cells associated with tumor stroma and vasculature. In this Prospect article we summarize evidence for SPARC and hevin in the regulation of tumor cell growth, differentiation, and metastasis, and we propose that matricellular proteins such as these perform critical functions in desmoplastic responses of tumors that culminate in their dissemination and eventual colonization of other sites.     

Revisiting the matricellular concept

The concept of a matricellular protein was first proposed by Paul Bornstein in the mid-1990s to account for the non-lethal phenotypes of mice with inactivated genes encoding thrombospondin-1, tenascin-C, or SPARC. It was also recognized that these extracellular matrix proteins were primarily counter or de-adhesive. This review reappraises the matricellular concept after nearly two decades of continuous investigation. The expanded matricellular family as well as the diverse and often unexpected functions, cellular location, and interacting partners/receptors of matricellular proteins are considered. Development of therapeutic strategies that target matricellular proteins are discussed in the context of pathology and regenerative medicine.     

Functional analysis of the matricellular protein SPARC with novel monoclonal antibodies.

SPARC (osteonectin, BM-40) is a matricellular glycoprotein that is expressed in many embryogenic and adult tissues undergoing remodeling or repair. SPARC modulates cellular interaction with the extracellular matrix (ECM), inhibits cell adhesion and proliferation, and regulates growth factor activity. To explore further the function and activity of this protein in tissue homeostasis, we have developed several monoclonal antibodies (MAbs) that recognize distinct epitopes on SPARC. The MAbs bind to SPARC with high affinity and identify SPARC by ELISA, Western blotting, immunoprecipitation, immunocytochemistry, and/or immunohistochemistry. The MAbs were also characterized in functional assays for potential alteration of SPARC activity. SPARC binds to collagen I and laminin-1 through an epitope defined by MAb 293; this epitope is not involved in the binding of SPARC to collagen III. The other MAbs did not interfere with the binding of SPARC to collagen I or III or laminin-1. Inhibition of the anti-adhesive effect of SPARC on endothelial cells by MAb 236 was also observed. Functional analysis of SPARC in the presence of these novel MAbs now confirms that the activities ascribed to this matricellular protein can be assigned to discrete subdomains.     

Molecular evolution of SPARC: absence of the acidic module and expression in the endoderm of the starlet sea anemone, Nematostella vectensis

The matricellular glycoprotein SPARC is composed of three functional domains that are evolutionarily conserved in organisms ranging from nematodes to mammals: a Ca²⁺-binding glutamic acid-rich acidic domain at the N-terminus (domain I), a follistatin-like module (domain II), and an extracellular Ca²⁺-binding (EC) module that contains two EF-hands and two collagen-binding epitopes (domain III). We report that four SPARC orthologs (designated nvSPARC1-4) are expressed by the genome of the starlet anemone Nematostella vectensis, a diploblastic basal cnidarian composed of an ectoderm and endoderm separated by collagen-based mesoglea. We also report that domain I is absent from all N. vectensis SPARC orthologs. In situ hybridization data indicate that N. vectensis SPARC mRNAs are restricted to the endoderm during post-gastrula development. The absence of the Ca²⁺-binding N-terminal domain in cnidarians and conservation of collagen-binding epitopes suggests that SPARC first evolved as a collagen-binding matricellular glycoprotein, an interaction likely to be dependent on the binding of Ca²⁺-ions to the two EF-hands in the EC domain. We propose that further Ca²⁺-dependent activities emerged with the acquisition of an acidic N-terminal module in triplobastic organisms.     

Enhanced growth of pancreatic tumors in SPARC-null mice is associated with decreased deposition of extracellular matrix and reduced tumor cell apoptosis

SPARC, a matricellular glycoprotein, modulates cellular interaction with the extracellular matrix (ECM). Tumor growth and metastasis occur in the context of the ECM, the levels and deposition of which are controlled in part by SPARC. Tumor-derived SPARC is reported to stimulate or retard tumor progression depending on the tumor type, whereas the function of host-derived SPARC in tumorigenesis has not been explored fully. To evaluate the function of endogenous SPARC, we have examined the growth of pancreatic tumors in SPARC-null (SP(-/-)) mice and their wild-type (SP(+/+)) counterparts. Mouse pancreatic adenocarcinoma cells injected s.c. grew significantly faster in SP(-/-) mice than cells injected into SP(+/+) animals, with mean tumor weights at sacrifice of 0.415 +/- 0.08 and 0.086 +/- 0.03 g (P < 0.01), respectively. Lack of endogenous SPARC resulted in decreased collagen deposition and fiber formation, alterations in the distribution of tumor-infiltrating macrophages, and decreased tumor cell apoptosis. There was no difference in microvessel density of tumors from SP(-/-) or SP(+/+) mice. However, tumors grown in SP(-/-) had a lower percentage of blood vessels that expressed smooth muscle alpha-actin, a marker of pericytes. These data reflect the importance of ECM deposition in regulating tumor growth and demonstrate that host-derived SPARC is a critical factor in the response of host tissue to tumorigenesis.     

SPARC regulates extracellular matrix organization through its modulation of integrin-linked kinase activity

SPARC, a 32-kDa matricellular glycoprotein, mediates interactions between cells and their extracellular matrix, and targeted deletion of Sparc results in compromised extracellular matrix in mice. Fibronectin matrix provides provisional tissue scaffolding during development and wound healing and is essential for the stabilization of mature extracellular matrix. Herein, we report that SPARC expression does not significantly affect fibronectin-induced cell spreading but enhances fibronectin-induced stress fiber formation and cell-mediated partial unfolding of fibronectin molecules, an essential process in fibronectin matrix assembly. By phage display, we identify integrin-linked kinase as a potential binding partner of SPARC and verify the interaction by co-immunoprecipitation and colocalization in vitro. Cells lacking SPARC exhibit diminished fibronectin-induced integrin-linked kinase activation and integrin-linked kinase-dependent cell-contractile signaling. Furthermore, induced expression of SPARC in SPARC-null fibroblasts restores fibronectin-induced integrin-linked kinase activation, downstream signaling, and fibronectin unfolding. These data further confirm the function of SPARC in extracellular matrix organization and identify a novel mechanism by which SPARC regulates extracellular matrix assembly.     

Cell cycle-dependent nuclear location of the matricellular protein SPARC: association with the nuclear matrix.

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular protein that inhibits cellular adhesion and proliferation. In this study, we report the detection of SPARC in the interphase nuclei of embryonic chicken cells in vivo. Differential partitioning of SPARC was also noted in the cytoplasm of these cells during discrete stages of M-phase: cells in metaphase and anaphase exhibited strong cytoplasmic immunoreactivity, whereas cells in telophase were devoid of labeling. Immunocytochemical analysis of embryonic chicken cells in vitro likewise showed the presence of SPARC in the nucleus. Furthermore, elution of soluble proteins and DNA from these cells indicated that SPARC might be a component of the nuclear matrix. We subsequently examined cultured bovine aortic endothelial cells, which initially appeared to express SPARC only in the cytoplasm. However, after elution of soluble proteins and chromatin, we also detected SPARC in the nuclear matrix of these cells. Embryonic chicken cells incubated with recombinant SPARC were seen to take up the protein and to translocate it to the nucleus progressively over a period of 17 h. These observations provide new information about SPARC, generally recognized as a secreted glycoprotein that mediates interactions between cells and components of the extracellular matrix. The evidence presented in this study indicates that SPARC might subserve analogous functions in the nuclear matrix.     

Thrombospondin-2 and SPARC/osteonectin are critical regulators of bone remodeling

Thrombospondin-2 (TSP2) and osteonectin/BM-40/SPARC are matricellular proteins that are highly expressed by bone cells. Mice deficient in either of these proteins show phenotypic alterations in the skeleton, and these phenotypes are most pronounced under conditions of altered bone remodeling. For example, TSP2-null mice have higher cortical bone volume and are resistant to bone loss associated with ovariectomy, whereas SPARC-null mice have decreased trabecular bone volume and fail to demonstrate an increase in bone mineral density in response to a bone-anabolic parathyroid hormone treatment regimen. In vitro, marrow stromal cell (MSC) osteoprogenitors from TSP2-null mice have increased proliferation but delayed formation of mineralized matrix. Similarly, in cultures of SPARC-null MSCs, osteoblastic differentiation and mineralized matrix formation are decreased. Overall, both TSP2 and SPARC positively influence osteoblastic differentiation. Intriguingly, both of these matricellular proteins appear to impact MSC fate through mechanisms that could involve the Notch signaling system. This review provides an overview of the role of TSP2 and SPARC in regulating bone structure, function, and remodeling, as determined by both in vitro and in vivo studies.     

Diverse biological functions of the SPARC family of proteins

The SPARC family of proteins represents a diverse group of proteins that modulate cell interaction with the extracellular milieu. The eight members of the SPARC protein family are modular in nature. Each shares a follistatin-like domain and an extracellular calcium binding E-F hand motif. In addition, each family member is secreted into the extracellular space. Some of the shared activities of this family include, regulation of extracellular matrix assembly and deposition, counter-adhesion, effects on extracellular protease activity, and modulation of growth factor/cytokine signaling pathways. Recently, several SPARC family members have been implicated in human disease pathogenesis. This review discusses recent advances in the understanding of the functional roles of the SPARC family of proteins in development and disease.     

Extracellular matrix remodeling in canine and mouse myocardial infarcts

Extracellular matrix proteins not only provide structural support, but also modulate cellular behavior by activating signaling pathways. Healing of myocardial infarcts is associated with dynamic changes in the composition of the extracellular matrix; these changes may play an important role in regulating cellular phenotype and gene expression. We examined the time course of extracellular matrix deposition in a canine and mouse model of reperfused infarction. In both models, myocardial infarction resulted in fragmentation and destruction of the cardiac extracellular matrix, extravasation of plasma proteins, such as fibrinogen and fibronectin, and formation of a fibrin-based provisional matrix providing the scaffold for the infiltration of granulation tissue cells. Lysis of the plasma-derived provisional matrix was followed by the formation of a cell-derived network of provisional matrix composed of cellular fibronectin, laminin, and hyaluronic acid and containing matricellular proteins, such as osteopontin and osteonectin/SPARC. Finally, collagen was deposited in the infarct, and the wound matured into a collagen-based scar with low cellular content. Although the canine and mouse infarcts exhibited a similar pattern of extracellular matrix deposition, deposition of the provisional matrix was more transient in the mouse infarct and was followed by earlier formation of a mature collagen-based scar after 7-14 days of reperfusion; at the same timepoint, the canine infarct was highly cellular and evolving. In addition, mature mouse infarcts showed limited collagen deposition and significant tissue loss leading to the formation of a thin scar. In contrast, dogs exhibited extensive collagen accumulation in the infarcted area. These species-specific differences in infarct wound healing should be taken into account when interpreting experimental infarction studies and when attempting to extrapolate the findings to the human pathological process.     

Expression of the matricellular protein SPARC in murine lens: SPARC is necessary for the structural integrity of the capsular basement membrane.

SPARC (Secreted Protein, Acidic and Rich in Cysteine) is a matricellular glycoprotein that modulates cell proliferation, adhesion, migration, and extracellular matrix (ECM) production. Although SPARC is generally abundant in embryonic tissues and is diminished in adults, we have found that the expression of SPARC in murine lens persists throughout embryogenesis and adulthood. Our previous studies showed that targeted ablation of the SPARC gene in mice results in cataract formation, a pathology attributed partially to an abnormal lens capsule. Here we provide evidence that SPARC is not a structural component of the lens capsule. In contrast, SPARC is abundant in lens epithelial cells, and newly differentiated fiber cells, with stable expression in wild-type mice up to 2 years of age. Pertubation of the lens capsule in animals lacking SPARC appears to be a consequence of the invasion of the lens cells situated beneath the capsule. Immunoreactivity for SPARC in the lens cells was uneven, with minimal reactivity in the epithelial cells immediately anterior to the equator. These epithelial cells appeared essentially noninvasive in SPARC-null mice, in comparison to the centrally located anterior epithelial cells, in which strong labeling by anti-SPARC IgG was observed. The posterior lens fibers exhibited cytoplasmic extensions into the posterior lens capsule, which was severely damaged in SPARC-null lenses. The expression of SPARC in wild-type lens cells, together with the abnormal lens capsule in SPARC-null mice, indicated that the structural integrity of the lens capsule is dependent on the matricellular protein SPARC. The effects of SPARC in the lens appear to involve regulation of lens epithelial and fiber cell morphology and functions rather than deposition as a structural component of the lens capsule.     

SPARC mediates early extracellular matrix remodeling following myocardial infarction

Secreted protein, acidic, and rich in cysteine (SPARC) is a matricellular protein that functions in the extracellular processing of newly synthesized collagen. Collagen deposition to form a scar is a key event following a myocardial infarction (MI). Because the roles of SPARC in the early post-MI setting have not been defined, we examined age-matched wild-type (WT; n=22) and SPARC-deficient (null; n=25) mice at day 3 post-MI. Day 0 WT (n=28) and null (n=20) mice served as controls. Infarct size was 52 ± 2% for WT and 47 ± 2% for SPARC null (P=NS), indicating that the MI injury was comparable in the two groups. By echocardiography, WT mice increased end-diastolic volumes from 45 ± 2 to 83 ± 5 μl (P < 0.05). SPARC null mice also increased end-diastolic volumes but to a lesser extent than WT (39 ± 3 to 63 ± 5 μl; P < 0.05 vs. day 0 controls and vs. WT day 3 MI). Ejection fraction fell post-MI in WT mice from 57 ± 2 to 19 ± 1%. The decrease in ejection fraction was attenuated in the absence of SPARC (65 ± 2 to 28 ± 2%). Fibroblasts isolated from SPARC null left ventricle (LV) showed differences in the expression of 22 genes encoding extracellular matrix and adhesion molecule genes, including fibronectin, connective tissue growth factor (CTGF; CCN2), matrix metalloproteinase-3 (MMP-3), and tissue inhibitor of metalloproteinase-2 (TIMP-2). The change in fibroblast gene expression levels was mirrored in tissue protein extracts for fibronectin, CTGF, and MMP-3 but not TIMP-2. Combined, the results of this study indicate that SPARC deletion preserves LV function at day 3 post-MI but may be detrimental for the long-term response due to impaired fibroblast activation.     

Enhanced angiogenesis characteristic of SPARC-null mice disappears with age

The impairment of angiogenesis in aging has been attributed, in part, to alterations in proteins associated with the extracellular matrix (ECM). SPARC (secreted protein acidic and rich in cysteine/osteonectin/BM-40) is a matricellular protein that regulates endothelial cell function as well as cell-ECM interactions. We have previously shown that angiogenesis, as reflected by fibrovascular invasion into subcutaneously implanted polyvinyl alcohol (PVA) sponges, is increased in SPARC-null mice (6-9 months of age) relative to their wild-type (WT) counterparts. In this study, we define the influence of aging on (a) the expression of SPARC and (b) fibrovascular invasion into sponge implants in SPARC-null and WT mice. The expression of SPARC in fibroblasts and endothelial cells derived from young donors (humans mean age less than 30 years and mice 4-6 months of age) and old donors (humans mean age over 65 years and mice 22-27 months of age) decreased 1.6 to 2.3-fold with age. Analysis of fibrovascular invasion into sponges implanted into old (22-27 months) SPARC-null and WT mice showed no differences in percent area of invasion or collagenous ECM. Moreover, sponges from old SPARC-null and WT mice contained similar levels of VEGF that were significantly lower than those from young (4-6 months) mice. In contrast to fibroblasts from young SPARC-null mice, dermal fibroblasts from old SPARC-null mice did not migrate farther, proliferate faster, or produce greater amounts of VEGF relative to their old WT counterparts. However, when stimulated with TGF-beta1, primary cells isolated from the sponge implants, and dermal fibroblasts from both old SPARC-null and WT mice, showed marked increases in VEGF secretion. These data indicate that aging results in a loss of enhanced angiogenesis in SPARC-null mice, as a result of the detrimental impact of age on cellular functions, collagen deposition, and VEGF synthesis. However, the influence of aging on these processes may be reversed, in part, by growth factor stimulation.     

Retention of the matricellular protein SPARC in the endoplasmic reticulum of chondrocytes from patients with pseudoachondroplasia.

Pseudoachondroplasia (PSACH) is an autosomal dominant disease characterized by dwarfism, morphological irregularities of long bones and hips, and early-onset osteoarthritis. This disease has been attributed to mutations in a structural protein of the cartilage extracellular matrix (ECM), cartilage oligomeric matrix protein (COMP), which result in its selective retention in the chondrocyte rough endoplasmic reticulum (ER). Accumulation of excessive amounts of mutated COMP might reflect a defect in protein trafficking by PSACH chondrocytes. Here we identify the matricellular protein SPARC as a component of this trafficking deficit. SPARC was localized to the hypertrophic chondrocytes in the normal human tibial growth plate and in cultured control cartilage nodules. In contrast, concentrated intracellular depots of SPARC were identified in nodules cultured from three PSACH patients with mutations in COMP. The accumulated SPARC was coincident with COMP and with protein disulfide isomerase, a resident chaperone of the rough ER, whereas SPARC and COMP were not coincident in the ECM of control or PSACH nodules. SPARC-null mice develop severe osteopenia and degenerative intervertebral disc disease, and exhibit attenuation of collagenous ECM. The retention of SPARC in the ER of chondrocytes producing mutant COMP indicates a new intracellular function for SPARC in the trafficking/secretion of cartilage ECM.