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Chondrosarcoma is a malignant bone tumor characterized by the production of a modified cartilage‑type extracellular matrix (ECM). In the present study, the expression levels of the small leucine‑rich proteoglycans (SLRPs), decorin, biglycan and lumican, were examined in the HTB94 human chondrosarcoma cell line. HTB94 cells were found to express and secrete the 3 SLRP members. RT‑qPCR and western blot analysis demonstrated that lumican was the most abundantly secreted SLRP, whereas decorin and biglycan expression levels were low. The utilization of short interfering RNA specific for the decorin, biglycan, and lumican genes resulted in the efficient downregulation of the respective mRNA levels (P≤0.001). The growth of the HTB94 cells was stimulated by lumican (P≤0.001), whereas their migration and adhesion were not affected (P=NS). By contrast, these cellular functions were not sensitive to a decrease in low endogenous levels of decorin and biglycan. Lumicandeficiency significantly inhibited both basal and insulin‑like growth factor I (IGF‑I)‑induced HTB94 cell growth (P≤0.001 andP≤0.01, respectively). These effects were executed through the insulin‑like growth factor I receptor (IGF‑IR), whose activation was markedly attenuated (P≤0.01) in lumican‑deficient HTB94 cells. The downregulation of lumican induced the substantial inhibition of extracellular regulated kinase (ERK1/2) activation (P≤ 0.01), indicating that ERK1/2 is a necessary component of lumican/IGF‑IR‑mediated HTB94 cell proliferation. Moreover, the lumican‑deficient cells exhibit increased mRNA levels of p53 (P≤0.05), suggesting that lumican facilitates HTB94 cell growth through an IGF‑IR/ERK1/2/p53 signaling cascade. On the whole, the findings of the present study demonstrate that endogenous lumican is a novel regulator of HTB94 cell growth.

Osteosarcoma (OS) is a mesenchymally derived, aggressive bone cancer. OS cells produce an aberrant nonmineralized or partly mineralized extracellular matrix (ECM) whose components participate in signaling pathways connected to specific pathogenic phenotypes of this bone cancer. The expression of biglycan (BGN), a secreted small leucine-rich proteoglycan (SLRP), is correlated to aggressive OS phenotype and resistance to chemotherapy. A constitutive signaling of IGF-IR signaling input in sarcoma progression has been established. Here, we show that biglycan activates the IGF-IR signaling pathway to promote MG63 biglycan-secreting OS cell growth by forming a complex with the receptor. Computational models of IGF-IR and biglycan docking suggest that biglycan binds IGF-IR dimer via its concave surface. Our binding free energy calculations indicate the formation of a stable complex. Biglycan binding results in prolonged IGF-IR activation leading to protracted IGF-IR-dependent cell growth response of the poorly-differentiated MG63 cells. Moreover, biglycan facilitates the internalization (p ≤ 0.01, p ≤ 0.001) and sumoylation-enhanced nuclear translocation of IGF-IR (p ≤ 0.05) and its DNA binding in MG63 cells (p ≤ 0.001). The tyrosine kinase activity of the receptor mediates this mechanism. Furthermore, biglycan downregulates the expression of the tumor-suppressor gene, PTEN (p ≤ 0.01), and increases the expression of endothelial–mesenchymal transition (EMT) and aggressiveness markers vimentin (p ≤ 0.01) and fibronectin (p ≤ 0.01) in MG63 cells. Interestingly, this mechanism is not valid in moderately and well-differentiated, biglycan non-expressing U-2OS and Saos-2 OS cells. Furthermore, biglycan exhibits protective effects against the chemotherapeutic drug, doxorubicin, in MG63 OS cells (p ≤ 0.01). In conclusion, these data indicate a potential direct and adjunct therapeutical role of biglycan in osteosarcoma.

Carcinogenesis is a multifactorial process with the input and interactions of environmental, genetic, and metabolic factors. During cancer development, a significant remodeling of the extracellular matrix (ECM) is evident. Proteoglycans (PGs), such as lumican, are glycosylated proteins that participate in the formation of the ECM and are established biological mediators. Notably, lumican is involved in cellular processes associated with tumorigeneses, such as EMT (epithelial-to-mesenchymal transition), cellular proliferation, migration, invasion, and adhesion. Furthermore, lumican is expressed in various cancer tissues and is reported to have a positive or negative correlation with tumor progression. This review focuses on significant advances achieved regardingthe role of lumican in the tumor biology. Here, the effects of lumican on cancer cell growth, invasion, motility, and metastasis are discussed, as well as the repercussions on autophagy and apoptosis. Finally, in light of the available data, novel roles for lumican as a cancer prognosis marker, chemoresistance regulator, and cancer therapy target are proposed.

The epithelial to mesenchymal transition (EMT) program is a crucial component in the processes of morphogenesis and embryonic development. The transition of epithelial to mesenchymal phenotype is associated with numerous structural and functional changes, including loss of cell polarity and tight cell-cell junctions, the acquisition of invasive abilities, and the expression of mesenchymal proteins. The switch between the two phenotypes is involved in human pathology and is crucial for cancer progression. Extracellular matrices (ECMs) are multi-component networks that surround cells in tissues. These networks are obligatory for cell survival, growth, and differentiation as well as tissue organization. Indeed, the ECM suprastructure, in addition to its supportive role, can process and deliver a plethora of signals to cells, which ultimately regulate their behavior. Importantly, the ECM derived signals are critically involved in the process of EMT during tumorigenesis. This review discusses the multilayer interaction between the ECM and the EMT process, focusing on contributions of discrete mediators, a strategy that may identify novel potential target molecules. Developmental Dynamics 247:368-381, 2018. © 2017 Wiley Periodicals, Inc.

The tumor microenvironment (TME) is composed of cancerous, non-cancerous, stromal, and immune cells that are surrounded by the components of the extracellular matrix (ECM). Glycosaminoglycans (GAGs), natural biomacromolecules, essential ECM, and cell membrane components are extensively altered in cancer tissues. During disease progression, the GAG fine structure changes in a manner associated with disease evolution. Thus, changes in the GAG sulfation pattern are immediately correlated to malignant transformation. Their molecular weight, distribution, composition, and fine modifications, including sulfation, exhibit distinct alterations during cancer development. GAGs and GAG-based molecules, due to their unique properties, are suggested as promising effectors for anticancer therapy. Considering their participation in tumorigenesis, their utilization in drug development has been the focus of both industry and academic research efforts. These efforts have been developing in two main directions; (i) utilizing GAGs as targets of therapeutic strategies and (ii) employing GAGs specificity and excellent physicochemical properties for targeted delivery of cancer therapeutics. This review will comprehensively discuss recent developments and the broad potential of GAG utilization for cancer therapy.

Osteosarcoma (OS) is a primary bone tumor of mesenchymal origin mostly affecting children and adolescents. The OS extracellular matrix (ECM) is extensively altered as compared to physiological bone tissue. Indeed, the main characteristic of the most common osteoblastic subtype of OS is non‑mineralized osteoid production. Parathyroid hormone (PTH) is a polypeptide hormone secreted by the chief cells of the parathyroid glands. The PTH-related peptide (PTHrP) may be comprised of 139, 141 or 173 amino acids and exhibits considerate N‑terminal amino acid sequence homology with PTH. The function of PTH/PTHrP is executed through the activation of the PTH receptor 1 (PTHR1) and respective downstream intracellular pathways which regulate skeletal development, bone turnover and mineral ion homeostasis. Both PTHR1 and its PTH/PTHrP ligands have been shown to be expressed in OS and to affect the functions of these tumor cells. This review aims to highlight the less well known aspects of PTH/PTHrP functions in the progression of OS by focusing on ECM-dependent signaling.

Hormone-dependent cancers exhibit high morbidity and mortality. In spite of advances in therapy, the treatment of hormone-dependent cancers remains an unmet health need. The tumor microenvironment (TME) exhibits unique characteristics that differ among various tumor types. It is composed of cancerous, non-cancerous, stromal, and immune cells that are surrounded and supported by components of the extracellular matrix (ECM). Therefore, the interactions among cancer cells, stromal cells, and components of the ECM determine cancer progression and response to therapy. Proteoglycans (PGs), hybrid molecules consisting of a protein core to which sulfated glycosaminoglycan chains are bound, are significant components of the ECM that are implicated in all phases of tumorigenesis. These molecules, secreted by both the stroma and cancer cells, are crucial signaling mediators that modulate the vital cellular pathways implicated in gene expression, phenotypic versatility, and response to therapy in specific tumor types. A plethora of deregulated signaling pathways contributes to the growth, dissemination, and angiogenesis of hormone-dependent cancers. Specific inputs from the endocrine and immune systems are some of the characteristics of hormone-dependent cancer pathogenesis. Importantly, the mechanisms involved in various aspects of cancer progression are executed in the ECM niche of the TME, and the PG components crucially mediate these processes. Here, we comprehensively discuss the mechanisms through which PGs affect the multifaceted aspects of hormone-dependent cancer development and progression, including cancer metastasis, angiogenesis, immunobiology, autophagy, and response to therapy.

Biglycan, a small leucine rich proteoglycan (SLRP), is an important participant in bone homeostasis and development as well as in bone pathology. In the present study biglycan was identified as a positive regulator of MG63 osteosarcoma cell growth (p ≤ 0.001). IGF-I was shown to increase biglycan expression (p ≤ 0.01), whereas biglycan-deficiency attenuated significantly both basal and IGF-I induced cell proliferation of MG63 cells (p ≤ 0.001; p ≤ 0.01, respectively). These effects were executed through the IGF-IR receptor whose activation was strongly attenuated (p ≤ 0.01) in biglycan-deficient MG63 cells. Biglycan, previously shown to regulate Wnt/β-catenin pathway, was demonstrated to induce a significant increase in β-catenin protein expression evident at cytoplasmic (p ≤ 0.01), membrane (p ≤ 0.01), and nucleus fractions in MG63 cells (p ≤ 0.05). As demonstrated by immunofluorescence, increase in β-catenin expression is attributed to co-localization of biglycan with the Wnt co-receptor low-density lipoprotein receptor-related protein 6 (LRP6) resulting in attenuated β-catenin degradation. Furthermore, applying anti-β-catenin and anti-pIGF-IR antibodies to MG-63 cells demonstrated a cytoplasmic and to the membrane interaction between these molecules that increased upon exogenous biglycan treatment. In parallel, the downregulation of biglycan significantly inhibited both basal and IGF-I-dependent ERK1/2 activation, (p ≤ 0.001). In summary, we report a novel mechanism where biglycan through a LRP6/β-catenin/IGF-IR signaling axis enhances osteosarcoma cell growth.

Proteoglycans (PGs), important constituents of the extracellular matrix, have been associated with cancer pathogenesis. Their unique structure consisting of a protein core and glycosaminoglycan chains endowed with fine modifications constitutes these molecules as capable cellular effectors important for homeostasis and contributing to disease progression. Indeed, differential expression of PGs and their interacting proteins has been characterized as specific for disease evolvement in various cancer types. Importantly, PGs to a large extent regulate the bioavailability of hormones, growth factors, and cytokines as well as the activation of their respective receptors which regulate phenotypic diversibility, gene expression and rates of recurrence in specific tumor types. Defining and targeting these effectors on an individual patient basis offers ground for the development of newer therapeutic approaches which may act as either supportive or a substitute treatment to the standard therapy protocols. This review discusses the roles of PGs in cancer progression, developing technologies utilized for the defining of the PG "signature" in disease, and how this may facilitate the generation of tailor-made cancer strategies.

The epithelial to mesenchymal transition (EMT) program is a crucial component in the processes of morphogenesis and embryonic development. The transition of epithelial to mesenchymal phenotype is associated with numerous structural and functional changes, including loss of cell polarity and tight cell-cell junctions, the acquisition of invasive abilities, and the expression of mesenchymal proteins. The switch between the two phenotypes is involved in human pathology and is crucial for cancer progression. Extracellular matrices (ECMs) are multi-component networks that surround cells in tissues. These networks are obligatory for cell survival, growth, and differentiation as well as tissue organization. Indeed, the ECM suprastructure, in addition to its supportive role, can process and deliver a plethora of signals to cells, which ultimately regulate their behavior. Importantly, the ECM derived signals are critically involved in the process of EMT during tumorigenesis. This review discusses the multilayer interaction between the ECM and the EMT process, focusing on contributions of discrete mediators, a strategy that may identify novel potential target molecules. Developmental Dynamics 247:368-381, 2018. © 2017 Wiley Periodicals, Inc.

Hyaluronan (HA), an unsulfated glycosaminoglycan, is an important component of the complex extracellular matrix network which surrounds and supports cells in tissues. HA is detected in all vertebrate tissues, but the bulk of HA is produced and deposited in the skin. In this review we focus on the role of HA in skin-associated inflammatory disease and wound healing. Properties of HA are directly dependent on its molecular weight. Thus, high molecular weight HA (HMWHA) is deposited in normal tissues during homeostasis and promotes their stability whereas low molecular weight HA fragments (LMWHA), on the other hand, may arise from enzymatic or chemical activities. The degradation of HMWHA to LMWHA fragments, often leads to the generation of biologically active oligosaccharides with different properties and postulated functions in wound scar formation and inflammation. More detailed studies of HA involvement in skin-associated inflammatory disease may result in novel treatment modalities.

Parathyroid hormone (PTH)(1-34), which has been established to have a dual effect on bone metabolism, was recently found to regulate osteosarcoma cell migration. A significant part of the bone anabolic action of PTH(1-34) is attributed to fibroblast growth factor (FGF)-2 stimulation. Furthermore, it was recently suggested that the FGF-proteoglycan axis may form an extracellular matrix-related regulatory feedback loop that controls osteoblastic lineage cell proliferation and execution of the osteogenic program. In this study, we investigated the possible participation of FGF-2 signaling in PTH(1-34)-dependent osteosarcoma cell migration. FGF-2 treatment of osteosarcoma cells resulted in a significant increase (P ≤ 0.01) in MG63 cell migration, similar to that caused by PTH(1-34). mRNA expression analysis of cells treated with PTH(1-34) showed a strong increase in FGF-2 transcript levels (P = 0.0015). Interestingly, the addition of FGF-2 to MG63 cells led to significant downregulation of small leucine-rich proteoglycan biglycan expression at both the mRNA (P ≤ 0.0001) and protein (60%) levels. In order to examine the significance of biglycan on MG63 cell migration, transfection with short interfering RNA specific for biglycan was performed, resulting in a significant increase (P ≤ 0.01) in the migration capacity of biglycan-deficient MG63 cells. In contrast, exogenous human recombinant biglycan strongly inhibited the migration of these cells (P ≤ 0.01). Finally, a direct correlation between PTH(1-34) action and biglycan expression was established by the finding of a significant decrease (P ≤ 0.01) in biglycan transcript levels in PTH(1-34)-treated cells. To summarize, the present study demonstrates a novel cooperative mechanism of PTH(1-34) and FGF-2 action that results in specific alteration of the biglycan extracellular matrix content to regulate osteosarcoma cell migration.

Estrogens are related with the growth and development of target tissues and play a critical role in breast cancer progression. The effects of estrogens are mediated by the estrogen receptors ERalpha and ERbeta, which are members of the nuclear steroid receptor superfamily. To date, it is not known how these hormones elicit many of their effects on extracellular matrix molecules and how these effects can be connected with ER expression. For this purpose, the effect of estradiol on ER expression as well as on proteoglycan and metalloproteinase expression was studied. The effect of E2 on extracellular matrix molecule expression has been studied using ERalpha suppression in breast cancer cells. Our studies using ERalpha-positive MCF-7 cells show that estradiol affects the expression of syndecan-2, but not of syndecan-4, through ERalpha. Furthermore, the ability of estradiol to affect MMP-9 and TIMP-1 expression is connected with ERalpha status. Together, these data demonstrate the significant role of ERalpha on mediating the effect of estradiol on extracellular matrix molecules.

Osteosarcoma is the most common primary bone tumour associated with childhood and adolescence. The possible role of the small leucine-rich proteoglycan, lumican, in the growth and metastasis of various cancer types has recently been investigated. In this study, the expression of lumican was examined in moderately differentiated (MG-63) and well-differentiated (Saos 2) human osteosarcoma cell lines of high and low metastatic capability, respectively. Real-time PCR, western blotting with antibodies against the protein core and keratan sulfate, and specific enzymatic digestions were the methods employed. The two human osteosarcoma cell lines were found to express and secrete lumican partly substituted with keratan sulfate glycosaminoglycans. Importantly, the non-metastatic, well-differentiated Saos 2 cells produced lumican at rates that were up to sevenfold higher than those of highly metastatic MG-63 cells. The utilization of short interfering RNA specific for the lumican gene resulted in efficient down-regulation of its mRNA levels in both cell lines. The growth of Saos 2 cells was inhibited by lumican, whereas their migration and chemotactic response to fibronectin were found to be promoted. Lumican expression was negatively correlated with the basal level of Smad 2 activation in these cells, suggesting that lumican may affect the bioavailability of Smad 2 activators. By contrast, these cellular functions of highly aggressive MG-63 cells were demonstrated not to be sensitive to a decrease in their low endogenous lumican levels. These results suggest that lumican expression may be positively correlated with the differentiation and negatively correlated with the progression of osteosarcoma.

Fibrosarcoma belongs to the sarcoma cancer group, which are spindle cell malignancies of mesenchymal origin, and owe their name to the predominant cell line that is present within the tumor. The extracellular matrix (ECM) is a complicated structure that surrounds and supports cells within tissues. Its main components are proteoglycans, collagens, glycoproteins, hyaluronan (HA), and several matrix-degrading enzymes. During cancer progression, significant changes can be observed in the structural and mechanical properties of ECM components. The ECM provides a physical scaffold to which tumor cells attach and migrate. Thus, it is required for key cellular events such as cell motility, adhesion, proliferation, invasion, and metastasis. Importantly, fibrosarcomas were shown to have a high content and turnover of ECM components including HA, proteoglycans, collagens, fibronectin, and laminin. In this review, we will focus on the HA component of fibrosarcoma ECM and critically discuss its role and involved mechanisms during fibrosarcoma pathogenesis.

Fibrosarcoma belongs to the sarcoma cancer group, which are spindle cell malignancies of mesenchymal origin, and owe their name to the predominant cell line that is present within the tumor. The extracellular matrix (ECM) is a complicated structure that surrounds and supports cells within tissues. Its main components are proteoglycans, collagens, glycoproteins, hyaluronan (HA), and several matrix-degrading enzymes. During cancer progression, significant changes can be observed in the structural and mechanical properties of ECM components. The ECM provides a physical scaffold to which tumor cells attach and migrate. Thus, it is required for key cellular events such as cell motility, adhesion, proliferation, invasion, and metastasis. Importantly, fibrosarcomas were shown to have a high content and turnover of ECM components including HA, proteoglycans, collagens, fibronectin, and laminin. In this review, we will focus on the HA component of fibrosarcoma ECM and critically discuss its role and involved mechanisms during fibrosarcoma pathogenesis.

Parathyroid hormone (PTH) strongly stimulates hyaluronan (HA) synthesis and secretion of both normal and carcinogenic cells of the osteoblastic lineage and improves skeletal microarchitecture. HA, a glycosaminoglycan component of the extracellular matrix (ECM), is capable of transmitting ECM-derived signals to regulate cellular function. In this study, we investigated whether the changes of HA metabolism induced by PTH (1–34) and PTH (7–84) peptides in moderately MG-63 and well-differentiated Saos 2 osteosarcoma cell lines, are correlated to their migration capabilities. Our results demonstrate that intermittent PTH (1–34) treatment significantly (P ≤ 0.01) supported the migration of MG-63 cells, increased their HA-synthase-2 (HAS2) expression (P ≤ 0.001), and enhanced their high-molecular size HA deposition in the pericellular matrix. Both increased endogenous HA production (P ≤ 0.01) and treatment with exogenous high-molecular weight HA (P ≤ 0.05) correlated to a significant increase of MG-63 cell migration capacity. Transfection with siHAS2 showed that PTH (1–34), mainly through HAS2, enhanced HA and regulated MG-63 cell motility. Interestingly, continuous PTH (1–34) treatment stimulated both Saos 2 cell HAS2 (P ≤ 0.001) and HAS1 (P ≤ 0.001) isoform expression inhibited their HYAL2 expression (P ≤ 0.001) and modestly (P ≤ 0.05) enhanced their migration. Therefore, the PTH (1–34) administration mode appears to distinctly modulate the migratory responses of the MG-63 moderately and Saos 2 well-differentiated osteosarcoma cell lines. Conclusively, the obtained data suggest that there is a regulatory effect of PTH (1–34), in an administration mode-dependent manner, on HA metabolism that is essential for osteosarcoma cell migration. © 2010 IUBMB IUBMB Life, 62(5): 377–386, 2010

Hyaluronan (HA) a glycosaminoglycan, is capable of transmitting extracellular matrix derived signals to regulate cellular functions. In this study, we investigated whether the changes in HT1080 and B6FS fibrosarcoma cell lines HA metabolism induced by basic fibroblast growth factor (bFGF) are correlated to their migration.Real-time PCR, in vitro wound healing assay, siRNA transfection, enzyme digestions, western blotting and immunofluorescence were utilized.bFGF inhibited the degradation of HA by decreasing hyaluronidase-2 expression in HT1080 cells (p=0.0028), increased HA-synthase-1 and -2 expression as we previously found and enhanced high molecular weight HA deposition in the pericellular matrix. Increased endogenous HA production (p=0.0022) and treatment with exogenous high molecular weight HA (p=0.0268) correlated with a significant decrease of HT1080 cell migration capacity. Transfection with siHAS2 and siHAS1 showed that mainly HAS1 synthesized high molecular weight HA regulates HT1080 cell motility. Induced degradation of the HA content by hyaluronidase treatment and addition of low molecular weight HA, resulted in a significant stimulation of HT1080 cells' motility (p0.01). In contrast, no effects on B6FS fibrosarcoma cell motility were observed.bFGF regulates, in a cell-specific manner the migration capability of fibrosarcoma cells by modulating their HA metabolism. HA metabolism is suggested to be a potential therapeutic target in fibrosarcoma.