Family Of Extracellular Matrix Proteins Research Articles

The biological functions and related signaling pathways of SPON2.

Spondin-2 (SPON2), also referred to as M-spondin or DIL-1, is a member of the extracellular matrix protein family known as Mindin-F-spondin (FS). SPON2 can be used as a broad-spectrum tumor marker for more than a dozen tumors, mainly prostate cancer. Meanwhile, SPON2 is also a potential biomarker for the diagnosis of certain non-tumor diseases. Additionally, SPON2 plays a pivotal role in regulating tumor metastasis and progression. In normal tissues, SPON2 has a variety of biological functions represented by promoting growth and development and cell proliferation. This paper presents a comprehensive overview of the regulatory mechanisms, diagnostic potential as a broad-spectrum biomarker, diverse biological functions, involvement in various signaling pathways, and clinical applications of SPON2.

C. elegans Hemidesmosomes Sense Collagen Damage to Trigger Innate Immune Response in the Epidermis.

The collagens are an enormous family of extracellular matrix proteins that play dominant roles in cell adhesion, migration and tissue remodeling under many physiological and pathological conditions. However, their function mechanisms in regulating innate immunity remain largely undiscovered. Here we use C. elegans epidermis as the model to address this question. The C. elegans epidermis is covered with a collagen-rich cuticle exoskeleton and can produce antimicrobial peptides (AMPs) against invading pathogens or physical injury. Through an RNAi screen against collagen-encoding genes, we found that except the previously reported six DPY collagens and the BLI-1 collagen, the majority of collagens tested appear unable to trigger epidermal immune defense when damaged. Further investigation suggests that the six DPY collagens form a specific substructure, which regulates the interaction between BLI-1 and the hemidesmosome receptor MUP-4. The separation of BLI-1 with MUP-4 caused by collagen damage leads to the detachment of the STAT transcription factor-like protein STA-2 from hemidesmosomes and the induction of AMPs. Our findings uncover the mechanism how collagens are organized into a damage sensor and how the epidermis senses collagen damage to mount an immune defense.

Editorial Highlights

Every organism is a model organism for understanding development, evolution, disease, and regeneration, and we have only begun to scratch the surface of the interdisciplinary genetic, molecular, cellular, and developmental mechanisms that regulate these biological processes. These “Highlights” denote exciting advances recently reported in Developmental Dynamics that illustrate the complex dynamics of developmental biology. Ocular Development and Disease. “Zebrafish model of RERE syndrome recapitulates key ophthalmic defects that are rescued by small molecule inhibitor of shh signaling” by Aman George, Jerry Lee, James Liu, Suzie Kim and Brian Brooks; DevDyn 252:4, pp. 495-509; https://doi.org/10.1002/dvdy.561. Mutations in the human RERE gene (ATROPHIN 2, OMIM *605226) are associated with a neuro-developmental disorder that includes ophthalmic, cardiac, genitourinary and hearing defects. This study demonstrates that rerea mutant (babyface) zebrafish recapitulate the optic fissure closure defects that result from loss of RERE function in humans. Interestingly, the RERE gene acts as a transcriptional corepressor in various nuclear receptor signaling pathways, and the optic fissure phenotype is associated with expansion of proximal retinal optic stalk in concert with deregulated shh signaling. Consistent with this model, inhibiting shh signaling can rescue the coloboma phenotype. Craniofacial Biology. “Frem1 activity is regulated by Sonic Hedgehog signaling in the cranial neural crest mesenchyme during midfacial morphogenesis” by Matthew McLaughlin, Miranda Sun, Tyler Beames, Austin Steward, Joshua Theisen, Hannah Chung, Joshua Everson, Ivan Moskowitz, Michael Sheets and Robert Lipinski; DevDyn 252:4, pp. 483-494; https://doi.org/10.1002/dvdy.555. The human face exhibits remarkable variation, and is also highly prone to developmental anomalies. The Frem/Fras family of extracellular matrix proteins has been implicated in regulating face shape variation as well as being mutated in the pathogenesis of disorders such as Manitoba-oculo-tricho-anal bifid nose with or without anorectal and renal anomalies, and Fraser syndromes. This study examined the activity of Frem1 during facial morphogenesis and identified Sonic Hedgehog (Shh) as an upstream regulator that promotes the proliferation of neural crest cells. Shh-Frem1 signaling is disrupted during the pathogenesis of midface hypoplasia, which provides deeper insights into the mechanisms underlying congenital craniofacial anomalies, and variation in midfacial morphogenesis. Organogenesis. “Foxd1 is required for 3D patterning of the kidney interstitial matrix by Sarah Lipp, Kathryn Jacobson, Andrew Schwaderer, David Hains and Sarah Calve; DevDyn 252:4, pp. 463-482; https://doi.org/10.1002/dvdy.545. The extracellular matrix (ECM) comprises a complex network of proteins and glycosaminoglycans that functions as a reservoir of growth factors, while also providing structural integrity and support during kidney development. This study combined 3D imaging and proteomics to investigate the role of Foxd1 in stromal cells, which are responsible for synthesizing the interstitial extracellular matrix, and document how the extracellular matrix dynamically changes during normal murine kidney development and in a model of kidney hypodysplasia. Changes in interstitial extracellular matrix corresponded to disruptions in stromal cell patterning, which helps to shed light on why human patients with connective tissue disorders have an increased incidence of kidney cysts and congenital anomalies of the kidney and urinary tract.

Structure, evolution and expression of zebrafish cartilage oligomeric matrix protein (COMP, TSP5). CRISPR-Cas mutants show a dominant phenotype in myosepta.

COMP (Cartilage Oligomeric Matrix Protein), also named thrombospondin-5, is a member of the thrombospondin family of extracellular matrix proteins. It is of clinical relevance, as in humans mutations in COMP lead to chondrodysplasias. The gene encoding zebrafish Comp is located on chromosome 11 in synteny with its mammalian orthologs. Zebrafish Comp has a domain structure identical to that of tetrapod COMP and shares 74% sequence similarity with murine COMP. Zebrafish comp is expressed from 5 hours post fertilization (hpf) on, while the protein is first detectable in somites of 11 hpf embryos. During development and in adults comp is strongly expressed in myosepta, craniofacial tendon and ligaments, around ribs and vertebra, but not in its name-giving tissue cartilage. As in mammals, zebrafish Comp forms pentamers. It is easily extracted from 5 days post fertilization (dpf) whole zebrafish. The lack of Comp expression in zebrafish cartilage implies that its cartilage function evolved recently in tetrapods. The expression in tendon and myosepta may indicate a more fundamental function, as in evolutionary distant Drosophila muscle-specific adhesion to tendon cells requires thrombospondin. A sequence encoding a calcium binding motif within the first TSP type-3 repeat of zebrafish Comp was targeted by CRISPR-Cas. The heterozygous and homozygous mutant Comp zebrafish displayed a patchy irregular Comp staining in 3 dpf myosepta, indicating a dominant phenotype. Electron microscopy revealed that the endoplasmic reticulum of myosepta fibroblasts is not affected in homozygous fish. The disorganized extracellular matrix may indicate that this mutation rather interferes with extracellular matrix assembly, similar to what is seen in a subgroup of chondrodysplasia patients. The early expression and easy detection of mutant Comp in zebrafish points to the potential of using the zebrafish model for large scale screening of small molecules that can improve secretion or function of disease-associated COMP mutants.

Repression of the PRELP gene is relieved by histone deacetylase inhibitors through acetylation of histone H2B lysine 5 in bladder cancer

BackgroundProline/arginine-rich end leucine-rich repeat protein (PRELP) is a member of the small leucine-rich proteoglycan family of extracellular matrix proteins, which is markedly suppressed in the majority of early-stage epithelial cancers and plays a role in regulating the epithelial–mesenchymal transition by altering cell–cell adhesion. Although PRELP is an important factor in the development and progression of bladder cancer, the mechanism of PRELP gene repression remains unclear.ResultsHere, we show that repression of PRELP mRNA expression in bladder cancer cells is alleviated by HDAC inhibitors (HDACi) through histone acetylation. Using ChIP-qPCR analysis, we found that acetylation of lysine residue 5 of histone H2B in the PRELP gene promoter region is a marker for the de-repression of PRELP expression.ConclusionsThese results suggest a mechanism through which HDACi may partially regulate the function of PRELP to suppress the development and progression of bladder cancer. Some HDACi are already in clinical use, and the findings of this study provide a mechanistic basis for further investigation of HDACi-based therapeutic strategies.

Comprehensive analysis of FRAS1/FREM family as potential biomarkers and therapeutic targets in renal clear cell carcinoma.

Background: FRAS1 (Fraser syndrome protein 1), together with FREM1 (the Fras1-related extracellular matrix proteins 1) and FREM2, belonging to the FRAS1/FREM extracellular matrix protein family, are considered to play essential roles in renal organogenesis and cancer progression. However, their roles in kidney renal clear cell carcinoma (KIRC) remain to be elucidated. Methods: FRAS1/FREM RNA expression analysis was performed using TCGA/GTEx databases, and valided using GEO databases and real-time PCR. Protein expression was peformed using CPTAC databases. Herein, we employed an array of bioinformatics methods and online databases to explore the potential oncogenic roles of FRAS1/FREM in KIRC. Results: We found that FRAS1, FREM1 and FREM2 genes and proteins expression levels were significantly decreased in KIRC tissues than in normal tissues. Decreased FRAS1/FREM expression levels were significantly associated with advanced clinicopathological parameters (pathological stage, grade and tumor metastasis status). Notably, the patients with decreased FRAS1/FREM2 expression showed a high propensity for metastasis and poor prognosis. FRAS1/FREM were correlated with various immune infiltrating cells, especially CD4+ T cells and its corresponding subsets (Th1, Th2, Tfh and Tregs). FRAS1 and FREM2 had association with DNA methylation and their single CpG methylation levels were associated with prognosis. Moreover, FRAS1/FREM might exert antitumor effects by functioning in key oncogenic signalling pathways and metabolic pathways. Drug sensitivity analysis indicated that high FRAS1 and FREM2 expression can be a reliable predictor of targeted therapeutic drug response, highlighting the potential as anticancer drug targets. Conclusion: Together, our results indicated that FRAS1/FREM family members could be potential therapeutic targets and valuable prognostic biomarkers of KIRC.

WNT1-inducible signalling pathway protein 1 stabilizes atherosclerotic plaques in apolipoprotein-E-deficient mice via the focal adhesion kinase/mitogen-activated extracellular signal-regulated kinase/extracellular signal-regulated kinase pathway.

The migration, proliferation and apoptosis of vascular smooth muscle cells (VSMCs) are critical for plaque stability. WNT-inducible signalling pathway protein-1 (WISP1), a member of the CCN family of extracellular matrix proteins, can expedite the migration and proliferation of VSMCs. However, its underlying mechanism and relationship with atherosclerosis remain elusive. The relationship between WISP1 and apoptosis of VSMCs has not been determined previously. In the study, we aimed to investigate the relationship between WISP1 and plaque stability and its related mechanism.ApoE-/- mice were divided following groups: the null lentivirus (NC), lentivirus WISP1 (IvWISP1) and WISP1-shRNA (shWISP1) groups. Immunofluorescence, Oil Red O and Masson's staining of the carotid arteries were performed. Transwell wound healing assay, CCK8 assay, and TdT-mediated dUTP nick-end labeling (TUNEL) staining were performed using VSMCs. The levels of WISP1, P38, C-Jun N-terminal kinase, extracellular signal-regulated kinase (ERK), mitogen-activated extracellular signal-regulated kinase (MEK), focal adhesion kinase (FAK), phosphatidylinositol 3-kinase (PI3K), Akt (also known as PKB, protein kinase B), mammalian target of rapamycin (mTOR), cleaved caspase3, Bcl2 and Bax were detected by western blotting. The relative area of lipids and monocytes/macrophages in the shWISP1 group increased compared with that of the NC group. However, the relative area of smooth muscle cell and collagen in the IvWISP1 group increased compared with that in the NC group. Therefore, WISP1 could stabilize atherosclerotic plaques. Besides, WISP1 accelerate the migration and proliferation of VSMCs via integrin α5β1 and FAK/MEK/ERK signalling pathways. In addition, WISP1 can inhibit the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway. WISP1 not only inhibits the apoptosis of VSMCs via the PI3K/Akt/mTOR pathway but also enhances the migration and proliferation of VSMCs via the integrin α5β1 and FAK/MEK/ERK pathways. Therefore, WISP1 could enhance the stability of atherosclerotic plaques.

Pericytes as mediators of infiltration of macrophages in multiple sclerosis

BackgroundMultiple sclerosis (MS) is a neurodegenerative condition of the central nervous system (CNS). It is associated with blood–brain barrier (BBB) breakdown and intravasation of leukocytes, particularly monocyte-derived macrophages, into the CNS. Pericytes are mural cells that are encased within the basement membrane of vasculature, and they contribute functionally to the neurovascular unit. These cells play an important role in maintaining BBB integrity and CNS homeostasis. However, the critical role of pericytes in mediating inflammation in MS or its models is unclear. Whether pericytes infiltrate into the CNS parenchyma in MS also needs clarification.MethodsCNS samples from the experimental autoimmune encephalomyelitis (EAE) mouse model of MS were collected at different time points for immunohistochemical analysis of pericytes along the inflamed vasculature. These findings were validated using MS brain specimens, and further analysis of pericyte involvement in inflammation was carried out by culturing primary pericytes and macrophages. Multiplex ELISA, transmigration assay and real-time PCR were used to study the inflammatory potential of pericytes in cultures.ResultsWe found that pericytes exhibit a heterogenous morphology, with notable elongation in the inflamed perivascular cuffs of EAE. This was manifested by a decrease in pericyte density but an increase in the coverage by pericytes along the vasculature. Chondroitin sulfate proteoglycans (CSPGs), a family of extracellular matrix proteins enriched within inflamed perivascular cuffs, elevated levels of pro-inflammatory chemokines/cytokines in pericytes in culture. Importantly, pericytes stimulated with CSPGs enhanced macrophage migration. We did not detect pericytes in the CNS parenchyma during EAE, and this was corroborated in MS brain samples.ConclusionsOur data suggest that pericytes seek to restore the BBB through increased coverage, but that their exposure to CSPGs prompt their facilitation of macrophages to enter the CNS to elevate neuroinflammation in EAE and MS.

CD157 and Brain Immune System in (Patho)physiological Conditions: Focus on Brain Plasticity.

Ectoenzyme and receptor BST-1/CD157 has been considered as a key molecule involved in the regulation of functional activity of cells in various tissues and organs. It is commonly accepted that CD157 catalyzes NAD+ hydrolysis and acts as a component of integrin adhesion receptor complex. Such properties are important for the regulatory role of CD157 in neuronal and glial cells: in addition to recently discovered role in the regulation of emotions, motor functions, and social behavior, CD157 might serve as an important component of innate immune reactions in the central nervous system. Activation of innate immune system in the brain occurs in response to infectious agents as well as in brain injury and neurodegeneration. As an example, in microglial cells, association of CD157 with CD11b/CD18 complex drives reactive gliosis and neuroinflammation evident in brain ischemia, chronic neurodegeneration, and aging. There are various non-substrate ligands of CD157 belonging to the family of extracellular matrix proteins (fibronectin, collagen I, finbrinogen, and laminin) whose activity is required for controlling cell adhesion and migration. Therefore, CD157 could control structural and functional integrity of the blood-brain barrier and barriergenesis. On the other hand, contribution of CD157 to the regulation of brain development is rather possible since in the embryonic brain, CD157 expression is very high, whereas in the adult brain, CD157 is expressed on neural stem cells and, presumably, is involved in the neurogenesis. Besides, CD157 could mediate astrocytes’ action on neural stem and progenitor cells within neurogenic niches. In this review we will summarize how CD157 may affect brain plasticity acting as a molecule at the crossroad of neurogenesis, cerebral angiogenesis, and immune regulation.

The Ig-like domain of Punctin/MADD-4 is the primary determinant for interaction with the ectodomain of neuroligin NLG-1

Punctin/MADD-4, a member of the ADAMTSL extracellular matrix protein family, was identified as an anterograde synaptic organizer in the nematode Caenorhabditis elegans. At GABAergic neuromuscular junctions, the short isoform MADD-4B binds the ectodomain of neuroligin NLG-1, itself a postsynaptic organizer of inhibitory synapses. To identify the molecular bases of their partnership, we generated recombinant forms of the two proteins and carried out a comprehensive biochemical and biophysical study of their interaction, complemented by an in vivo localization study. We show that spontaneous proteolysis of MADD-4B first generates a shorter N-MADD-4B form, which comprises four thrombospondin (TSP) domains and one Ig-like domain and binds NLG-1. A second processing event eliminates the C-terminal Ig-like domain along with the ability of N-MADD-4B to bind NLG-1. These data identify the Ig-like domain as the primary determinant for N-MADD-4B interaction with NLG-1 in vitro We further demonstrate in vivo that this Ig-like domain is essential, albeit not sufficient per se, for efficient recruitment of GABAA receptors at GABAergic synapses in C. elegans The interaction of N-MADD-4B with NLG-1 is also disrupted by heparin, used as a surrogate for the extracellular matrix component, heparan sulfate. High-affinity binding of heparin/heparan sulfate to the Ig-like domain may proceed from surface charge complementarity, as suggested by homology three-dimensional modeling. These data point to N-MADD-4B processing and cell-surface proteoglycan binding as two possible mechanisms to regulate the interaction between MADD-4B and NLG-1 at GABAergic synapses.

Analyzing the Effects of Peters Plus Syndrome Mutations on Human β1,3‐glucosyltransferase

Peters plus syndrome (PPS) is a severe autosomal recessive congenital disorder of glycosylation characterized by multiple structural anomalies. PPS patients have classic phenotypes, including Peters anomaly (an ocular anterior segment malformation), disproportionate short stature, brachydactyly, dysmorphic facial features, developmental delay, and variable other systemic abnormalities, including cleft lip/palate, and anomalies in heart, skeletal and genitourinary system. PPS patients have mutations in the β1,3‐glucosyltransferase (B3GLCT) gene, which encodes an enzyme that catalyzes the addition of a glucose to an O‐linked fucose on thrombospondin type‐1 repeats (TSRs), forming an unusual Glucoseβ1‐3Fucose (GlcFuc) disaccharide on TSRs. Protein O‐fucosyltransferase 2 (POFUT2) is the enzyme that adds the O‐fucose to a serine or threonine in the consensus sequence C1XX(S/T)C2 found in TSRs, where C1 and C2 are the first two of six conserved cysteines in TSRs. Currently 65 human proteins are predicted to contain TSRs, and 49 out of these proteins contain the consensus sequence for this GlcFuc disaccharide modification. Half of these proteins belong to ADAMTS/ADAMTS‐like family of extracellular matrix proteins and have critical roles in mediating angiogenesis, re‐structuring extracellular matrix, and developmental processes. Previous results demonstrate that secretion of several ADAMTS/L proteins, in particular ADAMTS20 TSR2‐8, is dependent on the presence of POFUT2 and B3GLCT. Since both POFUT2 and B3GLCT are localized in endoplasmic reticulum and only modify folded TSRs, this leads to the proposal that both POFUT2 and B3GLCT assist the folding of TSRs, modulating the secretion of TSR‐containing proteins. Many identified PPS mutations in B3GLCT are splicing mutations upstream of the catalytic domain and are predicted to result in non‐functional proteins. To analyze the missense and truncation PPS mutations downstream of the catalytic domain, we introduced these mutations separately into a plasmid encoding human B3GLCT and analyzed for their ability to rescue secretion of ADAMTS20 TSR2‐8 from B3GLCT‐null cells. Our results showed that these PPS missense and truncation mutants not only abolished their ability to rescue ADAMTS20 TSR2‐8 secretion, but also greatly reduced mutant protein levels compared to wild type B3GLCT within the cells. Mutant enzymes harboring mutations associated with Peters Plus Spectrum, which is a milder form of PPS with patients who have a subset of PPS phenotypes, all rescued ADAMTS20 TSR2‐8 secretion and had the same protein levels within the cells as wild type B3GLCT. On‐going experiments are comparing the enzymatic activity and protein stability of mutants versus wild type enzyme. Overall, by analyzing the impact of PPS and Peters Plus Spectrum mutations on B3GLCT, we are aiming to discover novel therapeutic targets and advancing the development of novel treatments to alleviate the symptoms of PPS as well as other congenital disorders of glycosylation. This project is supported by NIH grant HD096030.

Mammalian hemicentin 1 is assembled into tracks in the extracellular matrix of multiple tissues.

Hemicentins (HMCNs) are a family of extracellular matrix proteins first identified in Caenorhabditis elegans, with two orthologs (HMCN1 and 2) in vertebrates. In worms, HMCN is deposited at specific sites where it forms long, fine tracks that link two tissues by connecting adjacent basement membranes (BMs). By generating CRISPR/Cas9-mediated Hmcn1 and Hmcn2 knockout mice, we tested the hypothesis that HMCNs perform similar functions in mammals. Hmcn1 -/- mice were viable and fertile. Using new, knockout mouse-validated HMCN1 antibodies, HMCN1 was detected in wild-type mice as fine tracks along the BM of hair and whisker follicles, in the sclera of the eyes, and in the lumen of some lymphoid conduits. It was also observed in the mesangial matrix of the kidney glomerulus. However, HMCN1 deficiency did not affect the functions of these tissues, including adherence of coat hairs and whiskers, the sieving function of lymphoid conduits, or the immune response to injected antigens. HMCN2 deficiency did not lead to any discernible phenotypes on its own or when combined with HMCN1 deficiency. That Hmcn1 -/- , Hmcn2 -/- , and Hmcn1/2 double knockout mice did not display any overt phenotypes implicates compensation by other members of the fibulin family.

Pre-existing self-reactive IgA antibodies associated with primary graft dysfunction after lung transplantation

BackgroundPrimary graft Dysfunction (PGD) results in significant mortality and morbidity after lung transplantation (LT). The objective of this study was to evaluate if pre-existing antibodies to self-antigens in sera of LT recipients are associated with PGD. MethodsThe serum profiles of IgG and IgA autoantibodies were analyzed using a customized proteomic microarray bearing 124 autoantigens. Autoantibodies were analyzed using Mann-Whitney U test or Fisher exact test. The association of the autoantibodies with clinical phenotypes and survival was analyzed by Kaplan-Meier Survival Analysis. Receiver operating curve characteristics (ROC) were calculated to evaluate the predictive value of the autoantibodies for PGD. Results51 patients were included in this study. Autoantigen microarray analysis on the pre-transplantation samples identified 17 IgA and 3 IgG autoantibodies which were significantly higher in recipients who developed PGD compared to those who did not (adjusted p < .05 and fold change>1.5). 6 IgA Abs were significantly associated with survival. Taken as a panel, an elevation of 6 IgA Abs had significant predictive value for PGD. Area under the curve value for the panel was 0.9413 for PGD with ROC analysis. Notably, 6 of the 17 IgA autoantigen targets are belong to proteoglycan family of extracellular matrix proteins. ConclusionPre-existing IgG and IgA autoantibodies in LT patients correlate with PGD and with survival in a single center, small cohort of lung transplant recipients. Further validation is needed to confirm the findings in the study.

Mice Lacking the Matrilin Family of Extracellular Matrix Proteins Develop Mild Skeletal Abnormalities and Are Susceptible to Age-Associated Osteoarthritis.

Matrilins (MATN1, MATN2, MATN3 and MATN4) are adaptor proteins of the cartilage extracellular matrix (ECM), which bridge the collagen II and proteoglycan networks. In humans, dominant-negative mutations in MATN3 lead to various forms of mild chondrodysplasias. However, single or double matrilin knockout mice generated previously in our laboratory do not show an overt skeletal phenotype, suggesting compensation among the matrilin family members. The aim of our study was to establish a mouse line, which lacks all four matrilins and analyze the consequence of matrilin deficiency on endochondral bone formation and cartilage function. Matn1-4−/− mice were viable and fertile, and showed a lumbosacral transition phenotype characterized by the sacralization of the sixth lumbar vertebra. The development of the appendicular skeleton, the structure of the growth plate, chondrocyte differentiation, proliferation, and survival were normal in mutant mice. Biochemical analysis of knee cartilage demonstrated moderate alterations in the extractability of the binding partners of matrilins in Matn1-4−/− mice. Atomic force microscopy (AFM) revealed comparable compressive stiffness but higher collagen fiber diameters in the growth plate cartilage of quadruple mutant compared to wild-type mice. Importantly, Matn1-4−/− mice developed more severe spontaneous osteoarthritis at the age of 18 months, which was accompanied by changes in the biomechanical properties of the articular cartilage. Interestingly, Matn4−/− mice also developed age-associated osteoarthritis suggesting a crucial role of MATN4 in maintaining the stability of the articular cartilage. Collectively, our data provide evidence that matrilins are important to protect articular cartilage from deterioration and are involved in the specification of the vertebral column.

Decreased CCN3 in Systemic Sclerosis Endothelial Cells Contributes to Impaired Angiogenesis

Systemic sclerosis (SSc) is a rare and severe connective tissue disease combining autoimmune and vasculopathy features, ultimately leading to organ fibrosis. Impaired angiogenesis is an often silent and life-threatening complication of the disease. We hypothesize that CCN3, a member of the CCN family of extracellular matrix proteins, which is an antagonist of the profibrotic protein CCN2 as well as a proangiogenic factor, is implicated in SSc pathophysiology. We performed skin biopsies on 26 patients with SSc, both in fibrotic and nonfibrotic areas for 17 patients, and collected 18 healthy control skin specimens for immunohistochemistry and cell culture. Histological analysis of nonfibrotic and fibrotic SSc skin shows a systemic decrease of papillary dermis surface as well as disappearance of capillaries. CCN3 expression is systematically decreased in the dermis of patients with SSc compared with healthy controls, particularly in dermal blood vessels. Moreover, CCN3 is decreased invitro in endothelial cells from patients with SSc. We show that CCN3 is essential for endothelial cell migration and angiogenesis invitro. In conclusion, CCN3 may represent a promising therapeutic target for patients with SSc presenting with vascular involvement.

The Comparison Spondin 2 Levels in Primary Glomerular Diseases.

Spondin 2 (SPON2) plays an important role in multiple processes and is a member of the Spondin 2/F-spondin family of extracellular matrix proteins. We investigated serum SPON2 levels and its correlation with renal functions and urine protein excretion in different glomerular diseases. The cohort included 97 consecutive adults with persistant proteinuria (>300 mg/day) with the diagnosis of focal segmental glomerulosclerosis (FSGS), membranous glomerulonephritis (MN), IgA nephropathy (IgAN), membranoproliferative glomerulonephritis (MPGN), and AA amyloidosis and the control groups with 15 polycystic kidney disease (PKD) and 32 healthy people. Serum SPON2 levels in MN (64.6 ng/mL), FSGS (47.8 ng/mL), IgAN (52.6 ng/mL), MPGN (54.6 ng/mL), and AA amyloidosis (60.7 ng/mL) groups were higher than those of the control (26.4 ng/mL) and nonglomerular disease groups (PKD) (15.3 ng/mL). Only serum SPON2 levels were correlated with serum uric acid and triglyceride levels in patients with glomerular disease. This is the first study to show that serum SPON2 levels are similar in different glomerular diseases and that there is no correlation between SPON2 and proteinuria grade.

Genetic mutation of Frem3 does not causeFraser syndrome in mice.

QBRICK, FRAS1, and FREM2 compose a family of extracellular matrix proteins characterized by twelve consecutive CSPG repeats and single or multiple Calx-β motifs. Dysfunction of these proteins have been associated with Fraser syndrome, which is characterized by malformation of skin, eyes, digits, and kidneys. FREM3 is another member of the 12-CSPG protein family. However, it remains unknown whether genetic dysfunction of FREM3 also causes Fraser syndrome or another developmental disorder. Here we investigated a Frem3 mutant mouse line generated by CRISPR/Cas9-mediated genome editing. The FREM3 mutant homozygotes were born at the expected Mendelian ratio and did not possess any defects characteristic of Fraser syndrome. These results indicate that the dysfunction of FREM3 is not associated with Fraser syndrome.

The expression level of fibulin-2 in the circulating RNA (ctRNA) of epithelial tumor cells of peripheral blood and tumor tissue of patients with metastatic lung cancer.

The Fibulins are a recently discovered family of extracellular matrix proteins. In this study, expression levels of the fibulin-2 (FBLN2) gene and its role in the formation of different metastatic foci were investigated in lung cancer patients. We analyzed 106 lung cancer patients and eight paraffin-embedded tissues, and 27 ethnical-, age- and sex-matched healthy controls for expression levels of the FBLN2 gene. cDNAs obtained from the enriched epithelial cells of peripheral blood lymphocytes and tumor tissues of patients were amplified with specific primers for the target FBLN2 gene and HPRT1 housekeeping gene using quantitative real-time polymerase chain reaction. FBLN2 gene expression levels of the enriched epithelial cells of peripheral blood lymphocytes were found to be decreased approximately twofold in all subsets of patients compared to healthy controls. Our results indicate a significant difference between patient subgroups and controls [F(4.124) = 14.846, p0.05] among patient subgroups: bone metastases versus non-metastatic groups (p = 0.997), bone versus brain metastases (p = 0994), bone metastases versus two primary tumors (p = 0.999), brain metastases versus two primary tumors (p = 0.999), brain metastases versus non-metastatic (p = 0.755), non-metastatic versus two primary tumors (p = 0.996), non-metastatic versus all other metastatic patients (p = 0.731). Moreover, we found a 50-fold upregulation of FBLN2 gene expression in paraffin-embedded tissues compared with the enriched epithelial cells ofperipheral blood lymphocytes of patients. In the study, the enriched epithelial cells of peripheral blood lymphocytes of decreased FBLN2 expression was found to be correlated with metastasis. The fibulin-2 molecules might induce the metastatic potential through interaction with the other molecules in the microenvironment, nevertheless, it is needed further research whether the importance of FBLN2 on lung cancer oncogenesis and as a biomarker for metastatic lung cancer.

Cumulative prognostic power of laminin genes in colorectal cancer

BackgroundLaminins are a major family of extracellular matrix proteins and the main component of basement membranes. Laminins are involved in many if not all stages of cancer progression, and expression of laminin genes has prognostic value in various types of cancer, including colorectal. Only single laminin genes or components of a single laminin trimer with significant differential expression have been regarded as potential biomarkers to date.ResultsHere we compared prognostic power of classifiers constructed from sets of laminin genes with that of any single laminin gene. The analysis showed that cumulative prognostic power of sets of laminin genes was higher and was achieved already with pairs and triples of the genes. Interestingly, components of the pairs and the triples did not belong to any known laminin trimer, but, taken together with the gene weights, suggested higher LAMA4/LAMA5 expression ratio in patients with poor prognosis.ConclusionsAnalysis of the laminin expression profile rather than expression of the single genes or components of laminin trimers is useful for colorectal cancer prognosis in patients. High LAMA4/LAMA5 ratio is associated with increased permeability of basement membranes suggesting that basement membranes produced by colorectal tumors might be an important hindrance to their own dissemination in patients.

Thrombospondins and remodeling of the tumor microenvironment

Vascular remodeling defines cancer growth and aggressiveness. Although cancer cells produce pro-angiogenic signals, the fate of angiogenesis critically depends on the cancer microenvironment. Composition of the extracellular matrix (ECM) and tumor inflammation determine whether a cancer will remain dormant, will be recognized by the immune system and eliminated, or whether the tumor will develop and lead to the spread and metastasis of cancer cells. Thrombospondins (TSPs), a family of ECM proteins that has long been associated with the regulation of angiogenesis and cancer, regulate multiple physiological processes that determine cancer growth and spreading, from angiogenesis to inflammation, metabolic changes, and properties of ECM. Here, we sought to review publications that describe various functions of TSPs that link these proteins to regulation of cancer growth by modulating multiple physiological and pathological events that prevent or support tumor development. In addition to its direct effects on angiogenesis, TSPs have important roles in regulation of inflammation, immunity, ECM properties and composition, and glucose and insulin metabolism. Furthermore, TSPs have distinct roles as regulators of remodeling in tissues and tumors, such that the pathways activated by a single TSP can interact and influence each other. The complex nature of TSP interactions and functions, including their different cell- and tissue-specific effects, may lead to confusing results and controversial conclusions when taken out of the context of interdisciplinary and holistic approaches. However, studies of TSP functions and roles in different systems of the organism offer an integrative view of tumor remodeling and a potential for finding therapeutic targets that would modulate multiple complementary processes associated with cancer growth.