Small Leucine-rich Proteoglycan Research Articles

BiAb Mediated Restoration of the Linkage between Dystroglycan and Laminin-211 as a Therapeutic Approach for α-Dystroglycanopathies.

Patients with α-dystroglycanopathies, a subgroup of rare congenital muscular dystrophies, present with a spectrum of clinical manifestations that includes muscular dystrophy as well as CNS and ocular abnormalities. Although patients with α-dystroglycanopathies are genetically heterogeneous, they share a common defect of aberrant post-translational glycosylation modification of the dystroglycan alpha-subunit, which renders it defective in binding to several extracellular ligands such as laminin-211 in skeletal muscles, agrin in neuromuscular junctions, neurexin in the CNS, and pikachurin in the eye, leading to various symptoms. The genetic heterogeneity associated with the development of α-dystroglycanopathies poses significant challenges to developing a generalized treatment to address the spectrum of genetic defects. Here, we propose the development of a bispecific antibody (biAb) that functions as a surrogate molecular linker to reconnect laminin-211 and the dystroglycan beta-subunit to ameliorate sarcolemmal fragility, a primary pathology in patients with α-dystroglycan-related muscular dystrophies. We show that the treatment of LARGEmyd-3J mice, an α-dystroglycanopathy model, with the biAb improved muscle function and protected muscles from exercise-induced damage. These results demonstrate the viability of a biAb that binds to laminin-211 and dystroglycan simultaneously as a potential treatment for patients with α-dystroglycanopathy.

Osteoglycin (OGN) Inhibits Cell Proliferation and Invasiveness in Breast Cancer via PI3K/Akt/mTOR Signaling Pathway.

IntroductionPrevious studies have indicated that the small leucine-rich proteoglycan (SLR) osteoglycin (OGN) is downregulated in various cancers, including squamous cervical carcinoma, gastric cancer, and colorectal adenoma, indicating that OGN is a putative tumor suppressor. However, its exact role in the pathology of human cancers, especially breast cancer (BC), is not clear.MethodsThe expression of OGN in BC tissues was examined using qRT-PCR. Online databases were employed to analyze the correlation between OGN expression and clinicopathological characteristics. CCK-8 assay, colony formation assay, transwell migration and invasion assays were applied to detect cell proliferation, colony formation, migration and invasion of BC cells, respectively. Xenograft tumor models were constructed to explore the role of OGN on tumor growth in vivo.ResultsOGN expression was reduced in 24 paired BC samples compared with normal tissue. Decreased expression of OGN was correlated with greater pathological grade, a more aggressive tumor subtype, and poor overall survival. In vitro experiments showed that OGN overexpressed by plasmid transfection significantly inhibited cell proliferation, colony formation, migration, and invasion of BC cell lines. In xenograft tumor models, overexpression of OGN repressed the growth of MCF-7 cells in vivo and alleviated the compression of the tumor on surrounding structures. We also observed that OGN expression reversed EMT via repressing the PI3K/Akt/mTOR pathway.ConclusionThis study revealed that OGN could function as a tumor suppressor during breast carcinogenesis, and we contribute new evidence to the body of research on the SLRP family.

Osteoadherin serves roles in the regulation of apoptosis and growth in MC3T3‑E1 osteoblast cells.

Small leucine‑rich proteoglycans (SLRPs) are a class of proteoglycans that are characterized by small protein cores and structures of leucine‑rich repeats. SLRPs are expressed in most extracellular matrices and share numerous biological functions that are associated with binding of collagens and cell surface receptors. Osteoadherin (also termed osteomodulin) is encoded by the Omd gene and is a keratan sulfate proteoglycan of the class II subfamily of SLRPs. Osteoadherin is highly expressed in mineralized tissues, including bone and dentin; however, it's precise roles remain unknown. The present study determined the Omd expression levels and investigated the effects of over‑ and under‑expression of osteoadherin in osteoblastic cells. Omd mRNA expression increased with osteoblast differentiation in MC3T3‑E1 cells. In C2C12 cells, Omd mRNA expression was induced by bone morphogenetic protein (BMP)2. Reporter assays similarly demonstrated activation of the Omd gene promoter following co‑transfection with Smad1 and Smad4, which are intracellular signaling molecules of the BMP2 signaling pathway. Overexpression of Omd increased the viability and decreased caspase 3/7 activity in MC3T3‑E1 cells. By contrast, following transfection with small interfering RNA for Omd, viable cell numbers were decreased and caspase 3/7 activity was increased. Furthermore, overexpression of Omd reduced the expression of CCN family 2 in these cells. These results demonstrate that Omd expression is regulated during osteoblast differentiation, and that the protein product osteoadherin serves roles in the apoptosis and growth of osteoblast cells.

Multifaceted Roles of Asporin in Cancer: Current Understanding.

The small leucine-rich proteoglycan (SLRP) family consists of 18 members categorized into five distinct classes, the traditional classes I–III, and the non-canonical classes IV–V. Unlike the other class I SLRPs (decorin and biglycan), asporin contains a unique and conserved stretch of aspartate (D) residues in its N terminus, and germline polymorphisms in the D-repeat-length are associated with osteoarthritis and prostate cancer progression. Since the first discovery of asporin in 2001, previous studies have focused mainly on its roles in bone and joint diseases, including osteoarthritis, intervertebral disc degeneration and periodontal ligament mineralization. Recently, asporin gene expression was also reported to be dysregulated in tumor tissues of different types of cancer, and to act as oncogene in pancreatic, colorectal, gastric, and prostate cancers, and some types of breast cancer, though it is also reported to function as a tumor suppressor gene in triple-negative breast cancer. Furthermore, asporin is also positively or negatively correlated with tumor proliferation, migration, invasion, and patient prognosis through its regulation of different signaling pathways, including the TGF-β, EGFR, and CD44 pathways. In this review, we seek to elucidate the signaling pathways and functions regulated by asporin in different types of cancer and to highlight some important issues that require investigation in future research.

Decorin Regulates the Aggrecan Network Integrity and Biomechanical Functions of Cartilage Extracellular Matrix.

Joint biomechanical functions rely on the integrity of cartilage extracellular matrix. Understanding the molecular activities that govern cartilage matrix assembly is critical for developing effective cartilage regeneration strategies. This study elucidated the role of decorin, a small leucine-rich proteoglycan, in the structure and biomechanical functions of cartilage. In decorin-null cartilage, we discovered a substantial reduction of aggrecan content, the major proteoglycan of cartilage matrix, and mild changes in collagen fibril nanostructure. This loss of aggrecan resulted in significantly impaired biomechanical properties of cartilage, including decreased modulus, elevated hydraulic permeability, and reduced energy dissipation capabilities. At the cellular level, we found that decorin functions to increase the retention of aggrecan in the neo-matrix of chondrocytes, rather than to directly influence the biosynthesis of aggrecan. At the molecular level, we demonstrated that decorin significantly increases the adhesion between aggrecan and aggrecan molecules and between aggrecan molecules and collagen II fibrils. We hypothesize that decorin plays a crucial structural role in mediating the matrix integrity and biomechanical functions of cartilage by providing physical linkages to increase the adhesion and assembly of aggrecan molecules at the nanoscale.

Regulation of proteoglycan production by varying glucose concentrations controls fiber formation in tissue engineered menisci

Fibrillar collagens are highly prevalent in the extracellular matrix of all connective tissues and therefore commonly used as a biomaterial in tissue engineering applications. In the native environment, collagen fibers are arranged in a complex hierarchical structure that is often difficult to recreate in a tissue engineered construct. Small leucine rich proteoglycans as well as hyaluronan binding proteoglycans, aggrecan and versican, have been implicated in regulating fiber formation. In this study, we modified proteoglycan production in vitro by altering culture medium glucose concentrations (4500, 1000, 500, 250, and 125 mg/L), and evaluated its effect on the formation of collagen fibers inside tissue engineered meniscal constructs. Reduction of extracellular glucose resulted in a dose dependent decrease in total sulfated glycosaminoglycan (GAG) production, but minimal decreases of decorin and biglycan. However, fibromodulin doubled in production between 125 and 4500 mg/L glucose concentration. A peak in fiber formation was observed at 500 mg/L glucose concentration and corresponded with reductions in total GAG production. Fiber formation reduction at 125 and 250 mg/L glucose concentrations are likely due to changes in metabolic activity associated with a limited supply of glucose. These results point to proteoglycan production as a means to manipulate fiber architecture in tissue engineered constructs. Statement of significanceFibrillar collagens are highly prevalent in the extracellular matrix of all connective tissues; however achieving appropriate assembly and organization of collagen fibers in engineered connective tissues is a persistent challenge. Proteoglycans have been implicated in regulating collagen fiber organization both in vivo and in vitro, however little is known about methods to control proteoglycan production and the subsequent fiber organization in tissue engineered menisci. Here, we show that media glucose content can be optimized to control proteoglycan production and collagen fiber assembly, with optimal collagen fiber assembly occurring at sub-physiologic levels of glucose.

The "other" 15-40%: The Role of Non-Collagenous Extracellular Matrix Proteins and Minor Collagens in Tendon.

Extracellular matrix (ECM) determines the physiological function of all tissues, including musculoskeletal tissues. In tendon, ECM provides overall tissue architecture, which is tailored to match the biomechanical requirements of their physiological function, that is, force transmission from muscle to bone. Tendon ECM also constitutes the microenvironment that allows tendon-resident cells to maintain their phenotype and that transmits biomechanical forces from the macro-level to the micro-level. The structure and function of adult tendons is largely determined by the hierarchical organization of collagen type I fibrils. However, non-collagenous ECM proteins such as small leucine-rich proteoglycans (SLRPs), ADAMTS proteases, and cross-linking enzymes play critical roles in collagen fibrillogenesis and guide the hierarchical bundling of collagen fibrils into tendon fascicles. Other non-collagenous ECM proteins such as the less abundant collagens, fibrillins, or elastin, contribute to tendon formation or determine some of their biomechanical properties. The interfascicular matrix or endotenon and the outer layer of tendons, the epi- and paratenon, includes collagens and non-collagenous ECM proteins, but their function is less well understood. The ECM proteins in the epi- and paratenon may provide the appropriate microenvironment to maintain the identity of distinct tendon cell populations that are thought to play a role during repair processes after injury. The aim of this review is to provide an overview of the role of non-collagenous ECM proteins and less abundant collagens in tendon development and homeostasis. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:23-35, 2020.

Chemokine Pathway Can Be the Potential Therapeutic Target for Hypertrophic Scar

Hypertrophic scar (HTS) is a dermal form of fibroproliferative disease that develops after deep burns, skin injuries, and surgical wounds. HTS is defined as a visible, elevated scar that does not spread into surrounding tissues and often regresses. HTS often leads to physical and psychological issues for patients. Cellular and molecular agents, such as chemokines, bone marrow derived stem cells, TGF-β, fibroblast heterogeneity, toll-like receptor 4, and small leucine-rich proteoglycans, have been known to play an important role in wound healing and abnormal scar formation. Targeting these agents could have therapeutic potential for the treatment of pathological scars. HTS often causes contracture deformities, aesthetic disfigurement, and prolonged periods of hospitalisation and rehabilitation, leading to psychological complications. Numerous treatments have been described for HTS, but the optimal treatment has not yet been established. Chemokines are implicated in all stages of wound healing, but hyperactivity of these agents leads to excessive scarring. A better understanding of the mechanisms of chemokine action, such as the role of the signalling agents in wound healing and abnormal scar formation, will help to establish more effective therapeutic strategies for HTS.

Tumor-specific inhibitory action of decorin on different hepatoma cell lines

BackgroundIn spite of therapeutic approaches, liver cancer is still one of the deadliest type of tumor in which tumor microenvironment may play an active role in the outcome of the disease. Decorin, a small leucine-rich proteoglycan is not only responsible for assembly and maintenance of the integrity of the extracellular matrix, but a natural inhibitor of cell surface receptors, thus it exerts antitumorigenic effects. Here we addressed the question whether this effect of decorin is independent of the tumor phenotypes including differentiation, proliferation and invasion. MethodFour hepatoma cell lines HepG2, Hep3B, HuH7 and HLE, possessing different molecular backgrounds, were selected to investigate. After proliferation tests, pRTK arrays, WB analyses, and immunofluorescent examinations were performed on decorin treated and control cells for comparison. ResultsSignificant growth inhibitory potential of decorin on three out of four hepatoma cell lines was proven, however the mode of its action was different. Induction of p21WAF1/CIP1, increased inactivation of c-myc and β-catenin, and decrease of EGFR, GSK3β and ERK1/2 phosphorylation levels were observed in HepG2 cells, pathways already well-described in literature. However, in the p53 deficient Hep3B and HuH7, InsR and IGF-1R were the main receptors transmitting signals. In harmony with its receptor status, Hep3B cells displayed high level of activated AKT. As the cell line is retinoblastoma mutant, ATR/Chk1/Wee1 system might hinder the cell cycle in G2/M phase via phosphorylation of CDK1. In Huh7 cells, all RTKs were inhibited by decorin followed by downregulation of AKT. Furthermore, HuH7 cell line responded with concentration-dependent ERK activation and increased phospho-c-myc level. Decorin had only a non-significant effect on the proliferation rate of HLE cell line. However, it responded with a significant decrease of pAKT, c-myc and β-catenin activity. In this special cell line, the inhibition of TGFβ may be the first step of the protective effect of decorin. ConclusionsBased on our results decorin may be a candidate therapeutic agent in the battle against liver cancer, but several questions need to be answered. It is certain that decorin is capable to exert its suppressor effect in hepatoma cells without respect to their phenotype and molecular background.

Abstract 128: Effects of NSAIDs on gene expression of small leucine-rich proteoglycans in prostate cancer cells

Abstract Background: Malignant transformation has been shown to be favored by the interaction between cancer cells and the tumor microenvironment, with a special role attributed to molecular components present in both compartments, such as proteoglycans, including Small Leucine-Rich Proteoglycans (SLRPs). SLRPs are increasingly recognized as powerful signaling molecules involved in cancer growth and inflammation. Recent studies suggest that, among other cancers, prostate cancer (PCa) undergoes changes in the expression patterns of these components of the extracellular matrix to promote their survival, growth, and spread. Growing evidence links inflammation to initiation and progression to advanced metastatic disease of PCa and other human cancers. Non-steroidal anti-inflammatory drugs (NSAIDs) have been associated with reduced risk of several cancers, including PCa. The protective effect of NSAIDs has been attributed to inhibition of cancer metastasis. The aim of this work was to determine the expression profiles of SLRPs in a highly metastatic prostate cancer cell line (PC-3) compared to a non-tumorigenic cell line (PWR-1E) and to identify the effects of NSAIDs in the expression patterns of these molecular components. Methods: Differences in gene expression were determined by qRT-PCR for a panel of 15 SLRPs (ASPN, ECM2, LUM, PRELP, KERA, OMD, EPYC, OPTC, CHAD, NYX, PODN, OGN, BGN, FMOD, and DCN). Alterations in the expression of these genes in response to treatment with NSAIDs (ibuprofen and nimesulide) were also determined. Results: Compared to non-tumorigenic PWR-1E cells, highly metastatic PC-3 cells showed higher expression levels of seven genes encoding SLRPs (FMOD, BGN, LUM, ASPN, ECM2, PRELP and EPYC). Of the remaining eight genes, DCN was not differentially expressed while PODN, NYX, KERA, OMD, OPTC, OGN and CHAD had not detectable expression in either PC-3 or PWR-1E cells. Treatment of PC-3 cells with ibuprofen and nimesulide affected the expression pattern of genes encoding SLRPs. Ibuprofen treatment increased the expression of PRELP and BGN and decreased the expression of EPYC and ECM2. On the other hand, nimesulide treatment resulted in increased expression of PRELP and decreased expression of LUM, EPYC and ECM2. Conclusions: highly metastatic PC-3 cells had increased expression levels of SLRPs compared to non-tumorigenic PWR-1E cells. NSAIDs (Nimesulide and ibuprofen) changed the expression pattern of SLRPGs in metastatic PC-3 cells. Pharmacological effects of NSAIDs in prostate cancer warrants further investigation at the clinical settings. Additional studies are required to determine the clinical usefulness of NSAIDs in prostate cancer. Citation Format: Niradiz Reyes, Juan Rebollo, Jan Geliebter. Effects of NSAIDs on gene expression of small leucine-rich proteoglycans in prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 128.

Expression of minor cartilage collagens and small leucine rich proteoglycans may be relatively reduced in osteoarthritic cartilage

BackgroundIn osteoarthritis (OA), cartilage matrix is lost despite vigorous chondrocyte anabolism. In this study, we attempted to determine whether altered matrix synthesis is involved in this paradox in disease progression through gene expression analysis and ultrastructural analysis of collagen fibrils within the cartilage matrix.MethodsCartilage tissues were obtained from 29 end-stage OA knees and 11 control knees. First, cDNA microarray analysis was performed and the expression of 9 genes involved in collagen fibrillogenesis was compared between OA and control cartilages. Then their expression was investigated in further detail by a quantitative polymerase chain reaction (qPCR) analysis combined with laser capture microdissection. Finally, collagen fibril formation was compared between OA and control cartilage by transmission electron microscopy.ResultsThe result of the microarray analysis suggested that the expression of type IX and type XI collagens and fibrillogenesis-related small leucine-rich proteoglycans (SLRPs) may be reduced in OA cartilage relative to the type II collagen expression. The qPCR analysis confirmed these results and further indicated that the relative reduction in the minor collagen and SLRP expression may be more obvious in degenerated areas of OA cartilage. An ultrastructural analysis suggested that thicker collagen fibrils may be formed by OA chondrocytes possibly through reduction in the minor collagen and SLRP expression.ConclusionsThis may be the first study to report the possibility of altered collagen fibrillogenesis in OA cartilage. Disturbance in collagen fibril formation may be a previously unidentified mechanism underlying the loss of cartilage matrix in OA.

Class I and Class II small leucine-rich proteoglycans in human cutaneous pacinian corpuscles

Pacinian corpuscles are onion bulb-like multilayered mechanoreceptors that consist of a complicated structure of axon terminals, Schwann related cells (inner core), endoneural related cells (intermediate layer) and perineurial related cells (outer core-capsule). The cells forming those compartments are continuous and share the properties of that covering the nerve fibers. Small leucine-rich proteoglycans are major proteoglycans of the extracellular matrix and regulate collagen fibrillogenesis, cell signalling pathways and extracellular matrix assembly. Here we used immunohistochemistry to investigate the distribution of class I (biglycan, decorin, asporin, ECM2 and ECMX) and class II (fibromodulin, lumican, prolargin, keratocan and osteoadherin) small leucine-rich proteoglycans in human cutaneous Pacinian corpuscles. The distribution of these compounds was: the inner core express decorin, biglycan, lumican, fibromodulin, osteoadherin; the intermediate layer display immunoreactivity for osteoadherin; the outer core biglycan, decorin, lumican, fibromodulin and osteoadherin; and the capsule contains biglycan, decorin, fibromodulin, and lumican. Asporin, prolargin and keratocan were undetectable. These results complement our knowledge about the distribution of small leucine-rich proteoglycans in human Pacinian corpuscles, and help to understand the composition of the extracellular matrix in these sensory formations.

Significance of Tsukushi in lung cancer

ObjectivesTsukushi (TSK), a member of the small leucine-rich repeat proteoglycan (SLRP) family, plays multifunctional roles by interacting with signaling molecules during development. However, the role of TSK in cancer remains unknown. The aim of the present study was to investigate the biological significance of TSK in lung cancer. Materials and methodsImmunohistochemistry of lung cancer tissues and reverse transcription polymerase chain reaction (PCR) of lung cancer cell lines were carried out to detect TSK. Then, RNA sequence analysis, Gene Ontology analysis, quantitative real-time PCR, western blotting, cell counting assay, invasion assays, and xenograft studies were done in a human lung adenocarcinoma cell line, H1975 with modification of TSK expression levels, in order to investigate its biological roles, in particular epithelial–mesenchymal transition (EMT) and proliferation. ResultsTSK was found to be highly expressed in lung cancer tissues and cell lines. Modification of TSK expression levels in H1975 resulted in changes in molecules related to EMT, including cadherin-1, snail family transcriptional repressor 1, snail family transcriptional repressor 2, and vimentin. The results of cell counting and xenograft assays showed that TSK promotes cell proliferation. ConclusionsIn lung cancer cells, TSK is expressed more highly than the other SLRPs family members, and regulates the EMT and proliferation. Thus, TSK may be a key coordinator of multiple pathways and an important structural element in the lung cancer microenvironment.

Breaking down chronic inflammatory diseases: the role of biglycan in promoting a switch between inflammation and autophagy.

It is well established that biglycan, a small leucine-rich proteoglycan, acts as an extracellular matrix-derived danger signal in its soluble form. By binding to innate immunity Toll-like receptors (TLR) 2 and 4, biglycan initiates and perpetuates the inflammatory response. Previous work has conveyed that biglycan's role in inflammation extends far beyond its function as a canonical danger signal. It has been shown that biglycan acts in an anti-inflammatory capacity, wherein it tightly regulates the inflammatory response. In this review, we will discuss a paradigm shift to our understanding of biglycan signaling in inflammation. Mounting evidence suggests that the selective interactions between biglycan, TLRs, and their adapter proteins critically regulate downstream signaling and disease outcome. Biglycan can act as a high-affinity ligand for TLR coreceptors CD14 and CD44, further providing an additional layer of complexity. We propose a novel concept, that biglycan steers signaling toward inflammation by interacting with CD14, whereas it can trigger autophagy by binding to CD44. Thus, biglycan, and perhaps others soluble proteoglycans, could function as molecular switches which could either propagate the signaling of chronic inflammation or promote the resolution of inflammatory processes. Obviously, these new functions have broad implications in the regulation of various inflammatory diseases and could provide the basis for developing novel therapeutic regimens that would selectively target the interactions between biglycan, TLRs, coreceptors, and adapter molecules.

Leucine Rich Repeat Proteins: Sequences, Mutations, Structures and Diseases

Mutations in the genes encoding Leucine Rich Repeat (LRR) containing proteins are associated with over sixty human diseases; these include high myopia, mitochondrial encephalomyopathy, and Crohn's disease. These mutations occur frequently within the LRR domains and within the regions that shield the hydrophobic core of the LRR domain. The amino acid sequences of fifty-five LRR proteins have been published. They include Nod-Like Receptors (NLRs) such as NLRP1, NLRP3, NLRP14, and Nod-2, Small Leucine Rich Repeat Proteoglycans (SLRPs) such as keratocan, lumican, fibromodulin, PRELP, biglycan, and nyctalopin, and F-box/LRR-repeat proteins such as FBXL2, FBXL4, and FBXL12. For example, 363 missense mutations have been identified. Replacement of arginine, proline, or cysteine by another amino acid, or the reverse, is frequently observed. The diverse effects of the mutations are discussed based on the known structures of LRR proteins. These mutations influence protein folding, aggregation, oligomerization, stability, protein-ligand interactions, disulfide bond formation, and glycosylation. Most of the mutations cause loss of function and a few, gain of function.

Tumor-dependent Effects of Proteoglycans and Various Glycosaminoglycan Synthesizing Enzymes and Sulfotransferases on Patients' Outcome.

The small leucine-rich proteoglycans (SLRPs) biglycan (BGN) and decorin (DCN) linked with sulfated glycosaminoglycan (GAG) chains exhibit oncogenic or tumor suppressive potentials depending on the cellular context and association with GAGs. We hypothesized that structural alterations and expression levels of BGN, DCN and their associated chondroitin sulfate (CS) polymerizing enzymes, dermatan sulfate (DS) epimerases and various sulfatases might be correlated with the tumor (sub)type and patients' survival. We acquired breast cancer (BC) and glioma patients' datasets from cBioPortal and R2 Genomics. Structural alterations and the expression pattern of CS polymerizing enzymes, DS epimerases and carbohydrate sulfotransferases (CHST) were compared to that of BGN and DCN and correlated to their clinical relevance. In BC, no mutations, but amplifications (0.2 - 2.1 %) and deletions (0.05 - 0.4 %) were found in BGN, DCN and CS/DS enzymes. In contrast, missense and/or truncated mutations (0.1 - 0.5 %), but a reduced amplification rate (0 - 1.5 %) were found in glioma. When compared to BC, the structural abnormalities caused altered mRNA expression levels of BGN, DCN, GAG synthesizing enzymes and CHST. Mutations in SLPRs, CHSY1, CHST4 and CHSY3 were correlated with a poor prognosis in glioma, while lack of mutations and copy number variations in the SLRPs, CHSY3, CHST15 and DSE displayed an increased survival in BC. A distinct association of BGN and DCN with CHST, CS polymerizing enzymes and DS epimerases was found in BC and glioma. Thus, a unique pattern of structural alterations and expression, which has clinical relevance, was found for PGs and GAG synthesizing enzymes and CHST in BC and glioma, which might help to identify high-risk patients and to develop personalized therapeutics.

Biglycan evokes autophagy in macrophages via a novel CD44/Toll-like receptor 4 signaling axis in ischemia/reperfusion injury

Biglycan, a small leucine-rich proteoglycan, acts as a danger signal and is classically thought to promote macrophage recruitment via Toll-like receptors (TLR) 2 and 4. We have recently shown that biglycan signaling through TLR 2/4 and the CD14 co-receptor regulates inflammation, suggesting that TLR co-receptors may determine whether biglycan-TLR signaling is pro- or anti-inflammatory. Here, we sought to identify other co-receptors and characterize their impact on biglycan-TLR signaling. We found a marked increase in the number of autophagic macrophages in mice stably overexpressing soluble biglycan. Invitro, stimulation of murine macrophages with biglycan triggered autophagosome formation and enhanced the flux of autophagy markers. Soluble biglycan also promoted autophagy in human peripheral blood macrophages. Using macrophages from mice lacking TLR2 and/or TLR4, CD14, or CD44, we demonstrated that the pro-autophagy signal required TLR4 interaction with CD44, a receptor involved in adhesion, migration, lymphocyte activation, and angiogenesis. Invivo, transient overexpression of circulating biglycan at the onset of renal ischemia/reperfusion injury (IRI) enhanced M1 macrophage recruitment into the kidneys of Cd44+/+ and Cd44-/- mice but not Cd14-/- mice. The biglycan-CD44 interaction increased M1 autophagy and the number of renal M2 macrophages and reduced tubular damage following IRI. Thus, CD44 is a novel signaling co-receptor for biglycan, an interaction that is required for TLR4-CD44-dependent pro-autophagic activity in macrophages. Interfering with the interaction between biglycan and specific TLR co-receptors could represent a promising therapeutic intervention to curtail kidney inflammation and damage.

The role of osteomodulin on osteo/odontogenic differentiation in human dental pulp stem cells

BackgroundExtracellular matrix secretion and odontoblastic differentiation in human dental pulp stem cells (hDPSCs) are the cellular bases for reparative dentinogenesis. Osteomodulin (OMD) is a member of the small leucine-rich proteoglycan family distributed in the extracellular matrix but little is known about its role in osteo/odontogenic differentiation. The objective of this study was to investigate the role of OMD during osteo/odontoblastic differentiation of hDPSCs.MethodshDPSCs were selected using immune-magnetic beads and their capability of multi-differentiation was identified. OMD knockdown was achieved using short hairpin RNA (shRNA) lentivirus and was confirmed by western blot. Gene expression was measured by real-time qPCR and osteo/odontoblastic differentiation of hDPSCs was determined by alizarin red S staining.ResultsCompared with uninduced cells, the transcription of OMD was up-regulated by 35-fold at the late stage of osteo/odontogenic differentiation. shRNA-mediated gene silencing of OMD decreased the expression of odontoblastic genes, such as alkaline phosphatase (ALP), dentin matrix acidic phosphoprotein 1 (DMP1) and dentin sialophosphoprotein (DSPP). Besides, knockdown of OMD attenuated the mineralized nodules formation induced by osteo/odontogenic medium.ConclusionsThese results implied that OMD may play a pivotal role in modulating the osteo/odontoblastic differentiation of hDPSCs.

Collagen I-based scaffolds negatively impact fracture healing in a mouse-osteotomy-model although used routinely in research and clinical application

Although several biomaterials for bone regeneration have been developed in the last decades, clinical application of bone morphogenetic protein 2 is clinically only approved when applied on an absorbable bovine collagen I scaffold (ACS) (Helistat; ACS-H). In research, another ACS, namely Lyostypt (ACS-L) is frequently used as a scaffold in bone-linked studies. Nevertheless, until today, the influence of ACS alone on bone healing remains unknown. Unexpectedly, in vitro studies using ASC-H revealed a suppression of osteogenic differentiation and a significant reduction of cell vitality when compared to ASC-L. In mice, we observed a significant delay in bone healing when applying ACS-L in the fracture gap during femoral osteotomy. The results of our study show for the first time a negative influence of both ACS-H and ACS-L on bone formation demonstrating a substantial need for more sophisticated delivery systems for local stimulation of bone healing in both clinical application and research. Statement of SignificanceOur study provides evidence-based justification to promote the development and approval of more suitable and sophisticated delivery systems in bone healing research. Additionally, we stimulate researchers of the field to consider that the application of those scaffolds as a delivery system for new substances represents a delayed healing approach rather than a normal bone healing which could greatly impact the outcome of those studies and play a pivotal role in the translation to the clinics. Moreover, we provide impulses on underlying mechanism involving the roles of small-leucine rich proteoglycans (SLRP) for further detailed investigations.

Decorin antagonizes corneal fibroblast migration via caveolae-mediated endocytosis of epidermal growth factor receptor

Decorin (Dcn), a small leucine-rich proteoglycan, is involved in the regulation of corneal wound healing. Epidermal growth factor receptor (EGFR) plays a critical role in corneal fibroblasts proliferation, migration and extracellular matrix (ECM) modulation upon injury or infection. The present study aimed to investigate the mechanistic role of Dcn in EGFR internalization to the regulation of corneal stromal fibroblasts (CSFs) migration, a key step in the corneal wound healing. Human corneal stromal fibroblasts (hCSF) cultures were generated from donor corneas. At 70% confluence, cells were switched to serum-free conditions for 48 h and then treated with decorin (250 nM) in the presence or absence of EGF (100 ng/ml) for various time points (10–60 min). Cell lysates were subjected to proteome array analysis screening for 42 different phosphorylated human receptor tyrosine kinases (RTKs), immunocytochemistry, and western blots to analyze EGFR phosphorylation. The scratch-wound assay was performed to evaluate the effects of decorin on EGF-mediated hCSF migration. Dcn caused a rapid EGFR phosphorylation within 10 min of exposure in RTK blot defining its role as a biological ligand for EGFR in hCSFs. Prolonged exposure to Dcn caused complete disappearance of EGFR and inhibition of the hCSF migration in the scratch wound assay suggesting Dcn binding to EGFR causes EGFR down-regulation. Immunostaining studies indicated that Dcn-treatment to hCSFs internalizes Dcn-EGFR complex, which does not require tyrosine kinase activity when treated with the AG1478 inhibitor and co-localizes the complex to the perinuclear region. Next, we found that Dcn-EGFR complex does not follow canonical early endosome internalization as revealed by the EEA1 antibody instead binds to the CD63 antibody directed for degradation by the late endosome. We also found that Dcn regulates the EGFR recycling by preventing its binding to Rab11, a specific antibody for recycling endosome. Further, hCSFs-pretreated with pharmacological inhibitors, methyl-β-cyclodextrin and chlorpromazine and supplemented with Dcn suggested EGFR trafficking via the caveolae-mediated pathway. These results suggest that Dcn acts as a biological ligand for EGFR and modulates hCSF migration via EGFR down-regulation, thus playing a vital role in corneal wound healing.