Small leucine-rich proteoglycans inhibit CNS regeneration by modifying the structural and mechanical properties of the lesion environment.

Extracellular matrix (ECM) deposition after central nervous system (CNS) injury leads to inhibitory scarring in humans and other mammals, whereas it facilitates axon regeneration in the zebrafish. However, the molecular basis of these different fates is not understood. Here, we identify small leucine-rich proteoglycans (SLRPs) as a contributing factor to regeneration failure in mammals. We demonstrate that the SLRPs chondroadherin, fibromodulin, lumican, and prolargin are enriched in rodent and human but not zebrafish CNS lesions. Targeting SLRPs to the zebrafish injury ECM inhibits axon regeneration and functional recovery. Mechanistically, we find that SLRPs confer mechano-structural properties to the lesion environment that are adverse to axon growth. Our study reveals SLRPs as inhibitory ECM factors that impair axon regeneration by modifying tissue mechanics and structure, and identifies their enrichment as a feature of human brain and spinal cord lesions. These findings imply that SLRPs may be targets for therapeutic strategies to promote CNS regeneration.

Molecular cues for immune cells from small leucine-rich repeat proteoglycans in their extracellular matrix-associated and free forms.

In this review we highlight emerging immune regulatory functions of lumican, keratocan, fibromodulin, biglycan and decorin, which are members of the small leucine-rich proteoglycans (SLRP) of the extracellular matrix (ECM). These SLRPs have been studied extensively as collagen-fibril regulatory structural components of the skin, cornea, bone and cartilage in homeostasis. However, SLRPs released from a remodeling ECM, or synthesized by activated fibroblasts and immune cells contribute to an ECM-free pool in tissues and circulation, that may have a significant, but poorly understood foot print in inflammation and disease. Their molecular interactions and the signaling networks they influence also require investigations. Here we present studies on the leucine-rich repeat (LRR) motifs of SLRP core proteins, their evolutionary and functional relationships with other LRR pathogen recognition receptors, such as the toll-like receptors (TLRs) to bring some molecular clarity in the immune regulatory functions of SLRPs. We discuss molecular interactions of fragments and intact SLRPs, and how some of these interactions are likely modulated by glycosaminoglycan side chains. We integrate findings on molecular interactions of these SLRPs together with what is known about their presence in circulation and lymph nodes (LN), which are important sites of immune cell regulation. Recent bulk and single cell RNA sequencing studies have identified subsets of stromal reticular cells that express these SLRPs within LNs. An understanding of the cellular source, molecular interactions and signaling consequences will lead to a fundamental understanding of how SLRPs modulate immune responses, and to therapeutic tools based on these SLRPs in the future.

Involvement of small leucine-rich proteoglycans and telocytes in thin and thick human endometrium: immunohistochemical and ultrastructural examination.

Thin endometrium, defined as an endometrial thickness of less than 7 mm during the late follicular phase, is a common cause of frequent cancelation of embryo transfers or recurrent implantation failure during assisted reproductive treatment. Small proteoglycans regulate intracellular signaling cascades by bridging other matrix molecules and tissue elements, affecting cell proliferation, adhesion, migration, and cytokine concentration. The aim of the study is to investigate the role of small leucine-rich proteoglycans in the pathogenesis of thin and thick human endometrium and their differences from normal endometrium in terms of fine structure properties. Normal, thin, and thick endometrial samples were collected, and small leucine-rich proteoglycans (SLRPs), decorin, lumican, biglycan, and fibromodulin immunoreactivities were comparatively analyzed immunohistochemically. The data were compared statistically. Moreover, ultrastructural differences among the groups were evaluated by transmission electron microscopy. The immunoreactivities of decorin, lumican, and biglycan were higher in the thin endometrial glandular epithelium and stroma compared to the normal and thick endometrium (p < .001). Fibromodulin immunoreactivity was also higher in the thin endometrial glandular epithelium than in the normal and thick endometrium (p < .001). However, there was no statistical difference in the stroma among the groups. Ultrastructural features were not profoundly different among cases. Telocytes, however, were not seen in the thin endometrium in contrast to normal and thin endometrial tissues. These findings suggest a possible role of changes in proteoglycan levels in the pathogenesis of thin endometrium.

Small leucine rich proteoglycans: Biology, function and their therapeutic potential in the ocular surface.

Small leucine rich proteoglycans (SLRPs) are the largest family of proteoglycans, with 18 members that are subdivided into five classes. SLRPs are small in size and can be present in tissues as glycosylated and non-glycosylated proteins, and the most studied SLRPs include decorin, biglycan, lumican, keratocan and fibromodulin. SLRPs specifically bind to collagen fibrils, regulating collagen fibrillogenesis and the biomechanical properties of tissues, and are expressed at particularly high levels in fibrous tissues, such as the cornea. However, SLRPs are also very active components of the ECM, interacting with numerous growth factors, cytokines and cell surface receptors. Therefore, SLRPs regulate major cellular processes and have a central role in major fundamental biological processes, such as maintaining corneal homeostasis and transparency and regulating corneal wound healing. Over the years, mutations and/or altered expression of SLRPs have been associated with various corneal diseases, such as congenital stromal corneal dystrophy and cornea plana. Recently, there has been great interest in harnessing the various functions of SLRPs for therapeutic purposes. In this comprehensive review, we describe the structural features and the related functions of SLRPs, and how these affect the therapeutic potential of SLRPs, with special emphasis on the use of SLRPs for treating ocular surface pathologies.

Age-dependent mechanical properties of tail tendons in wild-type and mimecan gene-knockout mice - A preliminary study.

Mimecan, or osteoglycin, belongs to the family of small leucine-rich proteoglycans. In connective tissues mimecan is implicated in the development and maintenance of normal collagen fibrillar organization. Since collagen fibrils are responsible for tissue reinforcement, the absence of mimecan could lead to abnormal tissue mechanical properties. Here, we carried out a preliminary investigation of possible changes in the mechanical properties of tendons in mice lacking a functional mimecan gene, as a function of age. Tail tendons were dissected from mimecan gene knockout (KO) and wild type (WT) mice at ages 1, 4 and 8 months and mechanical properties evaluated using a microtensile testing equipment. Mimecan gene knockout resulted in changes in tendon elasticity- and fracture-related properties. While tendons of WT mice exhibited enhanced mechanical properties with increasing age, this trend was notably attenuated in mimecan KO tendons, with the exception of fracture strain. When genotype and age were considered as cross factors, the diminution in the mechanical properties of mimecan KO tendons was significant for yield strength, modulus and fracture strength. This effect appeared to affect the mice at 4 month old. These preliminary results suggest that mimecan may have a role in regulating age-dependent mechanical function in mouse tail tendon.

Loss of hepatic SMLR1 causes hepatosteatosis and protects against atherosclerosis due to decreased hepatic VLDL secretion.

BACKGROUND AND AIMS: The assembly and secretion of VLDL from the liver, a pathway that affects hepatic and plasma lipids, remains incompletely understood. We set out to identify players in the VLDL biogenesis pathway by identifying genes that are co-expressed with the MTTP gene that encodes for microsomal triglyceride transfer protein, key to the lipidation of apolipoprotein B, the core protein of VLDL. Using human and murine transcriptomic data sets, we identified small leucine-rich protein 1 ( SMLR1 ), encoding for small leucine-rich protein 1, a protein of unknown function that is exclusively expressed in liver and small intestine. APPROACH AND RESULTS: To assess the role of SMLR1 in the liver, we used somatic CRISPR/CRISPR-associated protein 9 gene editing to silence murine Smlr1 in hepatocytes ( Smlr1 -LKO). When fed a chow diet, male and female mice show hepatic steatosis, reduced plasma apolipoprotein B and triglycerides, and reduced VLDL secretion without affecting microsomal triglyceride transfer protein activity. Immunofluorescence studies show that SMLR1 is in the endoplasmic reticulum and Cis-Golgi complex. The loss of hepatic SMLR1 in female mice protects against diet-induced hyperlipidemia and atherosclerosis but causes NASH. On a high-fat, high-cholesterol diet, insulin and glucose tolerance tests did not reveal differences in male Smlr1 -LKO mice versus controls. CONCLUSIONS: We propose a role for SMLR1 in the trafficking of VLDL from the endoplasmic reticulum to the Cis-Golgi complex. While this study uncovers SMLR1 as a player in the VLDL assembly, trafficking, and secretion pathway, it also shows that NASH can occur with undisturbed glucose homeostasis and atheroprotection.

Osteoglycin: An ECM Factor Regulating Fibrosis and Tumorigenesis.

The extracellular matrix (ECM) is made up of noncellular components that have special properties for influencing cell behavior and tissue structure. Small leucine-rich proteoglycans (SLRPs) are nonfibrillar ECM components that serve as structural scaffolds and signaling molecules. osteoglycin (OGN), a class III SLRP, is a ubiquitous ECM component that not only helps to organize the extracellular matrix but also regulates a number of important biological processes. As a glycosylated protein in the ECM, OGN was originally considered to be involved in fiber assembly and was reported to have a connection with fibrosis. In addition to these functions, OGN is found in a variety of cancer tissues and is implicated in cellular processes linked to tumorigenesis, including cell proliferation, invasion, metastasis, and epithelial-mesenchymal transition (EMT). In this review, we summarize the structure and functions of OGN as well as its biological and clinical importance in the context of fibrotic illness and tumorigenesis. This review aims to improve our understanding of OGN and provide some new strategies for the treatment of fibrosis and cancer.

Mechanistic interrogation of mutation-independent disease modulators of RDEB identifies the small leucine-rich proteoglycan PRELP as a TGF-ß antagonist and inhibitor of fibrosis.

Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic extracellular matrix disease caused by deficiency in type VII collagen (Col VII). The disease manifests with devastating mucocutaneous fragility leading to progressive fibrosis and metastatic squamous cell carcinomas. Although Col VII abundance is considered the main predictor of symptom course, previous studies have revealed the existence of mutation-independent mechanisms that control disease progression. Here, to investigate and validate new molecular modifiers of wound healing and fibrosis in a natural human setting, and toward development of disease-modulating treatment of RDEB, we performed gene expression profiling of primary fibroblast from RDEB siblings with marked phenotypic variations, despite having equal COL7A1 genotype. Gene enrichment analysis suggested that severe RDEB was associated with enhanced response to TGF-ß stimulus, oxidoreductase activity, and cell contraction. Consistently, we found an increased response to TGF-ß, higher levels of basal and induced reactive oxygen species (ROS), and greater contractile ability in collagen lattices in RDEB fibroblasts (RDEBFs) from donors with severe RDEB vs mild RDEB. Treatment with antioxidants allowed a reduction of the pro-fibrotic and contractile phenotype. Importantly, our analyses revealed higher expression and deposition in skin of the relatively uncharacterized small leucine-rich extracellular proteoglycan PRELP/prolargin associated with milder RDEB manifestations. Mechanistic investigations showed that PRELP effectively attenuated fibroblasts' response to TGF-ß1 stimulus and cell contractile capacity. Moreover, PRELP overexpression in RDEBFs enhanced RDEB keratinocyte attachment to fibroblast-derived extracellular matrix in the absence of Col VII. Our results highlight the clinical relevance of pro-oxidant status and hyper-responsiveness to TGF-ß in RDEB severity and progression. Of note, our study also reveals PRELP as a novel and natural TGF-ß antagonist with a likely dermo-epidermal pro-adhesive capacity.

KLF9 and EPYC acting as feature genes for osteoarthritis and their association with immune infiltration.

BACKGROUND: Osteoarthritis, a common degenerative disease of articular cartilage, is characterized by degeneration of articular cartilage, changes in subchondral bone structure, and formation of osteophytes, with main clinical manifestations including increasingly serious swelling, pain, stiffness, deformity, and mobility deficits of the knee joints. With the advent of the big data era, the processing of mass data has evolved into a hot topic and gained a solid foundation from the steadily developed and improved machine learning algorithms. Aiming to provide a reference for the diagnosis and treatment of osteoarthritis, this paper using machine learning identifies the key feature genes of osteoarthritis and explores its relationship with immune infiltration, thereby revealing its pathogenesis at the molecular level. METHODS: From the GEO database, GSE55235 and GSE55457 data were derived as training sets and GSE98918 data as a validation set. Differential gene expressions of the training sets were analyzed, and the LASSO regression model and support vector machine model were established by applying machine learning algorithms. Moreover, their intersection genes were regarded as feature genes, the receiver operator characteristic (ROC) curve was drawn, and the results were verified using the validation set. In addition, the expression spectrum of osteoarthritis was analyzed by immunocyte infiltration and the co-expression correlation between feature genes and immunocytes was construed. CONCLUSION: EPYC and KLF9 can be viewed as feature genes for osteoarthritis. The silencing of EPYC and the overexpression of KLF9 are associated with the occurrence of osteoarthritis and immunocyte infiltration.

Osteoglycin (OGN) promotes tumorigenesis of pancreatic cancer cell via targeting ID4.

Pancreatic cancer (PC) is the seventh-leading cause of cancer-related mortality, and is associated with limited therapeutic options and poor prognosis. The extracellular matrix (ECM) represents the main component of the tumor microenvironment. Studies have found controversial roles of osteoglycin (OGN), a classical small leucine-rich proteoglycan found in the ECM in human malignancies; however, the significance of OGN in PC has not been determined. Here, the expression profiles of OGN in PC tissues and cell lines were evaluated by Gene Expression Profiling Interactive Analysis (GEPIA) database, immunohistochemistry, western blot, and quantitative PCR. OGN was found to be significantly upregulated in PC tissues and cell lines. Moreover, the expression of OGN was observed to be closely associated with TNM stage, stage III showed a higher OGN expression than that of stages I and II. Survival analysis showed that patients with PC showing high levels of OGN had low survival rates. The effects of OGN on cell proliferation and apoptosis were analyzed using MTT, CCK8, EdU and TUNEL assays. Wound-healing and invasion assays were conducted to test migratory and invasive abilities. Overexpression of OGN was demonstrated to promote proliferation, migration, and invasion, and inhibit apoptosis of PC cells. Further experiments revealed that inhibitor of DNA binding 4 (ID4) was upregulated by OGN. Silencing ID4 by small interfering RNA was shown to partially reverse the tumor-promoting effect of OGN. Collectively, our preliminary results indicate that the elevated expression of OGN may be associated with PC progression and may serve as a potential biomarker for the diagnosis and prognosis of PC. Targeting of OGN/ID4 axis may be a promising strategy in PC therapy.

Posterior Amorphous Corneal Dystrophy: New Chromosomal Breakpoints in the Small Leucine-Rich Proteoglycan-Coding Region.

PURPOSE: The purpose of this study was to report the clinical features and describe the results obtained by multimodal corneal imaging of a patient with novel chromosomal breakpoints of the 12q21.33 locus. METHODS: This study was a case report and literature review. RESULTS: A 12-year-old girl presented with visual loss whose examination revealed a best-corrected visual acuity of 20/50 in her right eye and 20/35 in her left eye and corneal flattening and gray sheet-like opacities deep in the stroma. Anterior segment optical coherence tomography and ultrabiomicroscopy showed an evenly distributed hyperreflective line in the posterior stroma. Confocal microscopy revealed enlarged keratocytes and the presence of small reflective deposits from the pre-Descemet line to the endothelium. In addition, a 447-kb deletion that included the small leucine-rich proteoglycan-coding region in locus 12q21.33 was found. She was, therefore, diagnosed with PACD. CONCLUSIONS: PACD is a rare genetic disorder of the cornea characterized by gray sheet-like opacification of the posterior stroma in combination with corneal flattening. Confocal microscopy provides histologic segmentation of each corneal layer and shows the degree to which they are affected. New chromosomal breakpoints of a deletion in the small leucine-rich proteoglycan-coding region are hereby reported. PACD may be a contiguous gene syndrome, and further tests are required to identify the exact position responsible for the phenotypic variation.

Small Leucine-Rich Proteoglycans in Tendon Wound Healing.

Significance: Tendon injury possesses a high morbidity rate and is difficult to achieve a satisfying prognosis with currently available treatment strategies. Current approaches used for tendon healing always lead to the formation of fibrovascular scar tissue, which significantly compromises the biomechanics of the healed tendon. Moreover, the related functional deficiency deteriorates over time with an increased injury recurrence risk. Small leucine-rich proteoglycans (SLRPs) link and interact with collagen fibrils to regulate tendon structure and biomechanics, which can provide a new and promising method in the field of tendon injury management. Recent Advances: The effect of SLRPs on tendon development has been extensively investigated. SLRP deficiency impairs tendon collagen fibril structure and biomechanic properties, while administration of SLRPs generally benefits tendon wound healing and regains better mechanical properties. Critical Issues: Current knowledge on the role of SLRPs in tendon development and regeneration mostly comes from uninjured knockout mice, and mainly focuses on the morphology description of collagen fibril profile and mechanical properties. Little is known about the regulatory mechanism on the molecular level. Future Directions: This article reviews the current knowledge in this highly translational topic and provides an evidence-based conclusion, thereby encouraging in-depth investigations of SLRPs in tendons and the development of SLRP-based treatments for desired tendon healing.

The role of SLRPs and large aggregating proteoglycans in collagen fibrillogenesis, extracellular matrix assembly, and mechanical function of fibrocartilage.

PURPOSE: Proteoglycans, especially small leucine rich proteoglycans (SLRPs), play major roles in facilitating the development and regulation of collagen fibers and other extracellular matrix components. However, their roles in fibrocartilage have not been widely reviewed. Here, we discuss both SLRP and large aggregating proteoglycan's roles in collagen fibrillogenesis and extracellular matrix assembly in fibrocartilage tissues such as the meniscus, annulus fibrosus (AF), and TMJ disc. We also discuss their expression levels throughout development, aging and degeneration, as well as repair. METHODS: A review of literature discussing proteoglycans and collagen fibrillogenesis in fibrocartilage was conducted and data from these manuscripts were analyzed and grouped to discuss trends throughout the tissue's architectural zones and developmental stage. RESULTS: The spatial collagen architecture of these fibrocartilaginous tissues is reflected in the distribution of proteoglycans expressed, suggesting that each proteoglycan plays an important role in the type of architecture presented and associated mechanical function. CONCLUSION: The unique structure-function relationship of fibrocartilage makes the varied architectures throughout the tissues imperative for their success and understanding the functions of these proteoglycans in developing and maintaining the fiber structure could inform future work in fibrocartilage replacement using tissue engineered constructs.

Novel regulatory roles of small leucine-rich proteoglycans in remodeling of the uterine cervix in pregnancy.

The cervix undergoes rapid and dramatic shifts in collagen and elastic fiber structure to achieve its disparate physiological roles of competence during pregnancy and compliance during birth. An understanding of the structure-function relationships of collagen and elastic fibers to maintain extracellular matrix (ECM) homeostasis requires an understanding of the mechanisms executed by non-structural ECM molecules. Small-leucine rich proteoglycans (SLRPs) play key functions in biology by affecting collagen fibrillogenesis and regulating enzyme and growth factor bioactivities. In the current study, we evaluated collagen and elastic fiber structure-function relationships in mouse cervices using mice with genetic ablation of decorin and/or biglycan genes as representative of Class I SLRPs, and lumican gene representative of Class II SLRP. We identified structural defects in collagen fibril and elastic fiber organization in nonpregnant mice lacking decorin, or biglycan or lumican with variable resolution of defects noted during pregnancy. The severity of collagen and elastic fiber defects was greater in nonpregnant mice lacking both decorin and biglycan and defects were maintained throughout pregnancy. Loss of biglycan alone reduced tissue extensibility in nonpregnant mice while loss of both decorin and biglycan manifested in decreased rupture stretch in late pregnancy. Collagen cross-link density was similar in the Class I SLRP null mice as compared to wild-type nonpregnant and pregnant controls. A broader range in collagen fibril diameter along with an increase in mean fibril spacing was observed in the mutant mice compared to wild-type controls. Collectively, these findings uncover functional redundancy and hierarchical roles of Class I and Class II SLRPs as key regulators of cervical ECM remodeling in pregnancy. These results expand our understating of the critical role SLRPs play to maintain ECM homeostasis in the cervix.

The Genes Encoding Small Leucine-Rich Proteoglycans Undergo Differential Expression Alterations in Colorectal Cancer, Depending on Tumor Location.

Small leucine-rich proteoglycans (SLRPs) regulate different processes and undergo significant alterations in various diseases. Colon carcinomas (CCs) are heterogeneous pathologies with important clinical and molecular differences depending on their location, which makes it interesting to analyze the alterations in SLRPs in right- and left-sided tumors (RS- and LSCCs). SLRP transcription levels were studied in 32 CCs using qPCR compared to healthy colon mucosae samples from the same patients, 20 of them from LSCCs and the remaining 12 from RSCCs. Protein expression of genes with significant differences in their transcriptions was analyzed by immunohistochemistry. The alterations observed were related to survival data. The arrangement of transcription of SLRPs was quite similar in ascending and descending colon, but RS- and LSCCs displayed different patterns of alteration, with a greater number of deregulations occurring in the latter. The analysis of protein expression also indicated changes in the location of these molecules, largely moving to the cell interior. While podocan underexpression showed a trend toward better outcomes, no differences were observed in terms of overall survival. In vitro studies using the HT29 tumor cell line suggest that deregulation of SLRPs could affect cell proliferation. SLRPs constitute new differential markers of RS- and LSCCs, showing differences dependent on the anatomical location of the tumor.

Novel corneal targeting cell penetrating peptide as an efficient nanocarrier with an effective antimicrobial activity.

Delivery of therapeutics to the ocular tissues is challenging due to various anatomical and physiological barriers imposed. Cell penetrating peptides (CPPs) have emerged as potent drug nanocarriers that have been shown to overcome these barriers and enhance bioavailability of therapeutic macromolecules in deep ocular tissues. In the present study, an ocular targeting CPP has been designed by exploring potential targets of anterior ocular tissues in particular receptors, transporters and glycosaminoglycans (GAGs). The novel 11 mer peptide sequence, Corneal Targeting Sequence 1 (CorTS 1), has been developed by modifying leucine rich repeat (LRR) motif ensuring that it interacts with small leucine rich proteoglycans and collagen present in the corneal stroma. CorTS 1 exhibited dose dependent cellular translocation from 5 µM in Human Corneal Epithelial cell line (HCE) with no cytotoxicity. CorTS 1 was also found to deliver protein cargo inside HCE cells. Ex vivo tissue penetration study of CorTS 1 demonstrated in goat eyes revealed an augmented accumulation of peptide in the stromal region of cornea than in aqueous humor. Interestingly, CorTS 1 showed an antimicrobial activity against MRSA and Fusarium dimerum. Therefore, CorTS 1 can be a promising candidate with dual traits of antimicrobial agent and nanocarrier for ocular drugs.

Small Leucine-Rich Proteoglycans (SLRPs) in the Retina.

Retinal diseases such as age-related macular degeneration (AMD), retinopathy of prematurity (ROP), and diabetic retinopathy (DR) are the leading causes of visual impairment worldwide. There is a critical need to understand the structural and cellular components that play a vital role in the pathophysiology of retinal diseases. One potential component is the family of structural proteins called small leucine-rich proteoglycans (SLRPs). SLRPs are crucial in many fundamental biological processes involved in the maintenance of retinal homeostasis. They are present within the extracellular matrix (ECM) of connective and vascular tissues and contribute to tissue organization and modulation of cell growth. They play a vital role in cell-matrix interactions in many upstream signaling pathways involved in fibrillogenesis and angiogenesis. In this comprehensive review, we describe the expression patterns and function of SLRPs in the retina, including Biglycan and Decorin from class I; Fibromodulin, Lumican, and a Proline/arginine-rich end leucine-rich repeat protein (PRELP) from class II; Opticin and Osteoglycin/Mimecan from class III; and Chondroadherin (CHAD), Tsukushi and Nyctalopin from class IV.

Biglycan: an emerging small leucine-rich proteoglycan (SLRP) marker and its clinicopathological significance.

Extracellular matrix (ECM) plays an important role in the structural organization of tissue and delivery of external cues to the cell. Biglycan, a class I small leucine-rich proteoglycans (SLRP), is a key component of the ECM that participates in scaffolding the collagen fibrils and mediates cell signaling. Dysregulation of biglycan expression can result in wide range of clinical conditions such as metabolic disorder, inflammatory disorder, musculoskeletal defects and malignancies. In this review, we aim to update our current understanding regarding the link between altered expression of biglycan and different clinicopathological states. Biglycan interacts with toll like receptors (TLR)-2 and TLR-4 on the immune cells which initiates inflammation and aggravates inflammatory disorders. ECM unbound soluble biglycan acts as a DAMP (danger associated molecular pattern) resulting in sterile inflammation. Dysregulation of biglycan expression is also observed in inflammatory metabolic conditions such as atherosclerosis and obesity. In cancer, high-biglycan expression facilitates tumor growth, invasion and metastasis which is associated with poor clinical outcome. As a pivotal structural component of the ECM, biglycan strengthens the musculoskeletal system and its absence is associated with musculoskeletal defects. Thus, SLRP biglycan is a potential marker which is significantly altered in different clinicopathological states.

Uncovering molecular targets for regenerative therapy in degenerative disc disease: do small leucine-rich proteoglycans hold the key?

BACKGROUND CONTEXT: Small leucine-rich proteoglycans (SLRPs) play an essential role in extracellular matrix (ECM) organization and function. Recently, dysregulation of SLRPs has been implicated in degenerative disc disease (DDD). An in-depth analysis using high-throughput proteomic sequencing might provide valuable information on their implications in health and disease. PURPOSE: To utilize proteomics for analyzing the expression of SLRPs in fetal, healthy adult, and degenerated discs, to identify possible molecular targets to halt or reverse the degenerative process. STUDY DESIGN: Experimental analysis. METHODS: Proteomic signatures of 8 magnetic resonance imaging (MRI) normal lumbar discs (ND) [harvested from brain dead alive organ donors] were compared to 8 fetal disc samples (FD) [harvested from fetal spines devoid of congenital anomalies following spontaneous or medical termination of pregnancy] and 8 degenerate discs (DD) [collected from patients undergoing fusion surgery]. The various functional pathways along with the differential expression of SLRPs and the associated changes in collagens, large proteoglycans (LLRPs), matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs) have been analyzed further using bioinformatics. This project was self-funded by the Ganga Orthopedic Research and Education Foundation. RESULTS: ESI-LC-MS/MS analysis revealed a total of 1,029 proteins in FD, 1,785 proteins in ND, and 1,775 proteins in DD. Fetal disc proteins were engaged mainly in ribosomal pathways (indicating active proliferation and regenerative potential). The healthy adult discs (ND) primarily participated in ECM maintenance and basic metabolic pathways, whereas the unique proteins of DD group were involved in inflammatory (Complement and coagulation cascades, Systemic Lupus Erythematosus and Leukocyte transendothelial migration) pathways and infective (Staphylococcus aureus infection, Prion diseases, Amoebiasis, Pertussis, and Legionellosis) channels which favor the recent concepts of inflammaging and subclinical infection as causes of DDD. Analysis of SLRPs revealed the upregulation of Biglycan in FDs and downregulation of Lumican, Decorin, Prolargin, and Chondroadherin in the DD group. The universal decrease in the abundance of SLRPs in the DD group was associated with an increase in MMPs and a reduction in TIMPs, collagen and LLRP content. CONCLUSIONS: Our study documents the influence of SLRPs in the maintenance of disc health and also the need for future research in using them for disc regeneration. CLINICAL SIGNIFICANCE: The various SLRPs that we identified are all known to have a beneficial influence on ECM integrity and a negative effect on the degenerative process at different stages in the evolution of degeneration. Biglycan, which is abundantly present in a fetus, may be suitable for regenerative therapy, and the other SLRPs like Lumican, Prolargin, Decorin, and Chondroadherin may serve the same purpose and/or as biomarkers.

The proteoglycan mimecan is associated with carotid plaque vulnerability and increased risk of future cardiovascular death.

BACKGROUND AND AIMS: A vulnerable plaque is an atherosclerotic plaque that is rupture-prone with a higher risk to cause cardiovascular symptoms such as myocardial infarction or stroke. Mimecan or osteoglycin is a small leucine-rich proteoglycan, important for collagen fibrillogenesis, that has been implicated in atherosclerotic disease, yet the role of mimecan in human atherosclerotic disease remains unknown. METHODS: 196 human atherosclerotic carotid plaques were immunostained for mimecan. Smooth muscle cells, macrophages and intraplaque haemorrhage were also measured with immunohistochemistry. Neutral lipids were stained with Oil Red O and calcium deposits were quantified. Plaque homogenate levels of MCP-1, IL-6 and MIP-1ß were measured using a Proximity Extension Assay and MMP-9 levels were measured using Mesoscale. Glycosaminoglycans, collagen and elastin were assessed by colorimetric assays and TGF-ß1, ß2 and ß3 were measured using a multiplex assay. Mimecan gene expression in THP-1 derived macrophages was quantified by qPCR and protein expression in vitro was visualized with immunofluorescence. Cardiovascular events were registered using medical charts and national registers during follow-up. RESULTS: Mimecan correlated positively with plaque area of lipids, macrophages, intraplaque haemorrhage and inversely with smooth muscle cell staining. Mimecan also correlated positively with plaque levels of MMP-9 and MCP-1. Mimecan was upregulated in THP-1 derived macrophages upon stimulation with MCP-1. Patients with high levels of mimecan (above median) had higher risk for cardiovascular death. CONCLUSIONS: This study indicates that mimecan is associated with a vulnerable plaque phenotype, possibly regulated by plaque inflammation. In line, plaque levels of mimecan independently predict future cardiovascular death.