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William B Stallcup - One of the best experts on this subject based on the ideXlab platform.
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The NG2 Proteoglycan in Pericyte Biology.
Advances in experimental medicine and biology, 2018Co-Authors: William B StallcupAbstract:Studies of pericytes have been retarded by the lack of appropriate markers for identification of these perivascular mural cells. Use of antibodies against the NG2 Proteoglycan as a pericyte marker has greatly facilitated recent studies of pericytes, emphasizing the intimate spatial relationship between pericytes and endothelial cells, allowing more accurate quantification of pericyte/endothelial cell ratios in different vascular beds, and revealing the participation of pericytes throughout all stages of blood vessel formation. The functional importance of NG2 in pericyte biology has been established via NG2 knockdown (in vitro) and knockout (in vivo) strategies that reveal significant deficits in blood vessel formation when NG2 is absent from pericytes. NG2 influences pericyte proliferation and motility by acting as an auxiliary receptor that enhances signaling through integrins and receptor tyrosine kinase growth factor receptors. By acting in a trans orientation, NG2 also activates integrin signaling in closely apposed endothelial cells, leading to enhanced maturation and formation of endothelial cell junctions. NG2 null mice exhibit reduced growth of both mammary and brain tumors that can be traced to deficits in tumor vascularization. Use of Cre-Lox technology to produce pericyte-specific NG2 null mice has revealed specific deficits in tumor vessels that include decreased pericyte ensheathment of endothelial cells, diminished assembly of the vascular basement membrane, reduced vessel patency, and increased vessel leakiness. Interestingly, myeloid-specific NG2 null mice exhibit even larger deficits in tumor vascularization, leading to correspondingly slower tumor growth. Myeloid-specific NG2 null mice are deficient in their ability to recruit macrophages to tumors and other sites of inflammation. This absence of macrophages deprives pericytes of a signal that is crucial for their ability to interact with endothelial cells. The interplay between pericytes, endothelial cells, and macrophages promises to be an extremely fertile area of future study.
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distinct NG2 Proteoglycan dependent roles of resident microglia and bone marrow derived macrophages during myelin damage and repair
PLOS ONE, 2017Co-Authors: Karolina Kucharova, William B StallcupAbstract:We used a bone marrow transplantation approach to distinguish the activities of bone marrow-derived macrophages from the activities of central nervous system-resident microglia in phenomena associated with axon demyelination and remyelination. We transplanted wild type or germline NG2 null beta-actin-EGFP expressing bone marrow into irradiated wild type or NG2 null recipient mice, followed by analysis of lysolecithin-induced spinal cord demyelination and remyelination and quantification of Iba-1+/ F4/80+/ EGFP+ macrophages and Iba-1+/ F4/80+/ EGFP- microglia. One week after microinjection of 1% lysolecithin into the spinal cord, wild type recipients receiving NG2 null bone marrow exhibit greatly reduced infiltration of macrophages into lesions, compared to wild type recipients receiving wild type bone marrow. Wild type bone marrow recipients also exhibit larger numbers of demyelinated axons than NG2 null recipients, indicative of macrophage participation in the initial myelin damage. However, wild type bone marrow recipients also exhibit superior myelin repair at 6 weeks post-injury, compared to NG2 null bone marrow recipients, demonstrating the additional importance of macrophages in remyelination. Incompletely repaired lesions in NG2 null bone marrow recipients at 6 weeks post-injury retain elevated numbers of macrophages, in contrast to lower numbers of macrophages in more completely repaired lesions in wild type bone marrow recipients. This suggests that NG2 expression renders macrophages more effective in myelin repair and less likely to promote chronic inflammation. Effective macrophage involvement in myelin repair is due in part to effects on the proliferation and/or recruitment of oligodendrocyte progenitor cells. Reduced numbers of oligodendrocyte progenitors are seen in lesions in NG2 null bone marrow recipients, likely due to deficits in macrophage production of oligodendrocyte progenitor-relevant mitogens and in phagocytosis of inhibitory myelin debris. Microglia also appear to be important for clearance of myelin debris, as indicated by reduced phagocytosis in NG2 null recipients receiving wild type bone marrow.
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NG2 Proteoglycan enhances brain tumor progression by promoting beta 1 integrin activation in both cis and trans orientations
Cancers, 2017Co-Authors: William B StallcupAbstract:By physically interacting with beta-1 integrins, the NG2 Proteoglycan enhances activation of the integrin heterodimers. In glioma cells, co-localization of NG2 and 31 integrin in individual cells (cis interaction) can be demonstrated by immunolabeling, and the NG2-integrin interaction can be confirmed by co-immunoprecipitation. NG2-dependent integrin activation is detected via use of conformationally sensitive monoclonal antibodies that reveal the activated state of the beta-1 subunit in NG2-positive versus NG2-negative cells. NG2-dependent activation of beta-1 integrins triggers downstream activation of FAK and PI3K/Akt signaling, resulting in increased glioma cell proliferation, motility, and survival. Similar NG2-dependent cis activation of beta-1 integrins occurs in microvascular pericytes, leading to enhanced proliferation and motility of these vascular cells. Surprisingly, pericyte NG2 is also able to promote beta-1 integrin activation in closely apposed endothelial cells (trans interaction). Enhanced beta-1 signaling in endothelial cells promotes endothelial maturation by inducing the formation of endothelial junctions, resulting in increased barrier function of the endothelium and increased basal lamina assembly. NG2-dependent beta-1 integrin signaling is therefore important for tumor progression by virtue of its affects not only on the tumor cells themselves, but also on the maturation and function of tumor blood vessels.
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NG2 Proteoglycan dependent contributions of pericytes and macrophages to brain tumor vascularization and progression
Microcirculation, 2016Co-Authors: William B Stallcup, Weonkyoo You, Fusanori Yotsumoto, Karolina Kucharova, Pilar CejudomartinAbstract:The NG2 Proteoglycan promotes tumor growth as a component of both tumor and stromal cells. Using intracranial, NG2-negative B16F10 melanomas, we have investigated the importance of PC and Mac NG2 in brain tumor progression. Reduced melanoma growth in Mac-NG2ko and PC-NG2ko mice demonstrates the importance of NG2 in both stromal compartments. In each genotype, the loss of PC-endothelial cell interaction diminishes the formation of endothelial junctions and assembly of the basal lamina. Tumor vessels in Mac-NG2ko mice have smaller diameters, reduced patency, and increased leakiness compared to PC-NG2ko mice, thus decreasing tumor blood supply and increasing hypoxia. While the reduced PC interaction with endothelial cells in PC-NG2ko mice results from the loss of PC activation of β1 integrin signaling in endothelial cells, reduced PC-endothelial cell interaction in Mac-NG2ko mice results from 90% reduced Mac recruitment. The absence of Mac-derived signals in Mac-NG2ko mice causes the loss of PC association with endothelial cells. Reduced Mac recruitment may be due to diminished activation of integrins in the absence of NG2, causing decreased Mac interaction with endothelial adhesion molecules that are needed for extravasation. These results reflect the complex interplay that occurs between Mac, PC, and endothelial cells during tumor vascularization.
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NG2 Proteoglycan ablation reduces foam cell formation and atherogenesis via decreased low density lipoprotein retention by synthetic smooth muscle cells
Arteriosclerosis Thrombosis and Vascular Biology, 2016Co-Authors: Zhigang She, Yunchao Chang, Hongbo Pang, Wenlong Han, Houzao Chen, Jeffrey W Smith, William B StallcupAbstract:Objectives— Obesity and hyperlipidemia are critical risk factors for atherosclerosis. Because ablation of NG2 Proteoglycan in mice leads to hyperlipidemia and obesity, we investigated the impact of NG2 ablation on atherosclerosis in apoE null mice. Approach and Results— Immunostaining indicates that NG2 expression in plaque, primarily by synthetic smooth muscle cells, increases during atherogenesis. NG2 ablation unexpectedly results in decreased (30%) plaque development, despite aggravated obesity and hyperlipidemia. Mechanistic studies reveal that NG2-positive plaque synthetic smooth muscle cells in culture can sequester low-density lipoprotein to enhance foam-cell formation, processes in which NG2 itself plays direct roles. In agreement with these observations, low-density lipoprotein retention and lipid accumulation in the NG2/ApoE knockout aorta is 30% less than that seen in the control aorta. Conclusions— These results indicate that synthetic smooth muscle cell–dependent low-density lipoprotein retention and foam cell formation outweigh obesity and hyperlipidemia in promoting mouse atherogenesis. Our study sheds new light on the role of synthetic smooth muscle cells during atherogenesis. Blocking plaque NG2 or altering synthetic smooth muscle cells function may be promising therapeutic strategies for atherosclerosis.
Akiko Nishiyama - One of the best experts on this subject based on the ideXlab platform.
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NG2 cells properties progeny and origin
Brain Research Reviews, 2010Co-Authors: Jacqueline Trotter, Khalad Karram, Akiko NishiyamaAbstract:Abstract The NG2 Proteoglycan is a type 1-transmembrane protein expressed by a range of cell types within and outside the mammalian nervous system. NG2-expressing (NG2) cells are found in grey and white matter tracts of the developing and adult CNS and have previously been assumed to represent oligodendrocyte precursor cells: new work using transgenic mice has shown that NG2 cells generate oligodendrocytes, protoplasmic astrocytes and in some instances neurons in vivo. NG2 cells express GABAA receptors and the AMPA subtype of glutamate receptors. They make intimate contact to neurons prior to myelinating axons and also form electron-dense synaptic specialisations with axons in the cerebellum, cortex and hippocampus and with non-myelinated axons in the corpus callosum. These synaptic NG2 cells respond to neuronal release of glutamate and GABA. This neuron–glia interaction may thus regulate the differentiation and proliferation of NG2 cells. The C-terminal PDZ-binding motif of the NG2 protein binds several PDZ proteins including Mupp1, Syntenin and the Glutamate Receptor Interacting Protein (GRIP). Since GRIP can bind subunits of the AMPA receptors expressed by NG2 cells, the interaction between GRIP and NG2 may orientate the glial AMPA receptors towards sites of neuronal glutamate release. The origin, heterogeneity and function of NG2 cells as modulators of the neuronal network are important incompletely resolved questions.
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two separate metalloproteinase activities are responsible for the shedding and processing of the NG2 Proteoglycan in vitro
Molecular and Cellular Neuroscience, 2005Co-Authors: Richard A Asher, Joel M. Levine, Akiko Nishiyama, Daniel A Morgenstern, Francesca Properzi, James W FawcettAbstract:A high proportion of NG2 in the adult rat spinal cord is saline-soluble and migrates slightly faster than intact NG2 on SDS-PAGE, suggesting that it represents the shed ectodomain of NG2. In the injured cerebral cortex, much of the overall increase in NG2 is due to the saline-soluble (shed), rather than the detergent-soluble (intact), form. Hydroxamic acid metalloproteinase inhibitors, but not TIMPs, were able to prevent NG2 shedding in oligodendrocyte precursor cells (OPCs) in vitro. The generation of another truncated form of NG2 was, however, sensitive to TIMP-2 and TIMP-3. Two observations suggest that NG2 is involved in PDGF signaling in OPCs: the rate of NG2 shedding increased with cell density and NG2 expression was increased in the absence of PDGF. Ectodomain shedding converts NG2 into a diffusible entity able to interact with the growth cone, and we suggest that this release is likely to enhance its axon growth-inhibitory activity.
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transient expression of the NG2 Proteoglycan by a subpopulation of activated macrophages in an excitotoxic hippocampal lesion
Glia, 2001Co-Authors: Nadia Akhtar, Akiko NishiyamaAbstract:Cells that express the NG2 Proteoglycan (NG2+ cells) constitute a large glial population in the normal mature rodent brain. They can differentiate into oligodendrocytes but are distinct from mature oligodendrocytes, astrocytes, microglia, and neurons. Changes in NG2+ cells were examined in kainic acid-induced excitotoxic lesions of the hippocampus, and the relationship between NG2+ cells and reactive astrocytes and microglia was investigated between 1 and 90 days after lesioning. Two types of reactive NG2+ cells with altered morphology and increased NG2 immunoreactivity were observed in the lesion. Early changes, consisting of an increase in NG2 immunoreactivity and the number of processes, were apparent 24 h after lesioning and persisted through 3 months. These cells were distinct from reactive astrocytes or activated microglia/macrophages. A second type of reactive NG2+ cells appeared 2 weeks after injection, following an influx of macrophages. They had large, round cell bodies with short processes and expressed the microglia/macrophage antigens OX42 and ED1. Single cells coexpressing NG2 and macrophage/microglial antigens could be isolated from the lesion. The number of NG2+/OX42+ cells gradually declined and disappeared by 3 months after injection. They did not express glial fibrillary acidic protein or the alpha receptor for platelet-derived growth factor, indicating that they are distinct from astrocytes or oligodendrocyte progenitor cells. Cells that coexpressed NG2 and OX42 were never observed in hippocampal slice cultures treated with kainic acid, suggesting that NG2+/OX42+ cells are not derived from endogenous resident brain cells. These findings demonstrate that NG2 expression is transiently upregulated on activated macrophages/microglia that appear during the chronic stage in an excitotoxic lesion in the adult CNS.
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cytoskeletal reorganization induced by engagement of the NG2 Proteoglycan leads to cell spreading and migration
Molecular Biology of the Cell, 1999Co-Authors: Xuexun Fang, Akiko Nishiyama, Kimberlee Dahlinhuppe, Michael A Burg, Diana Barritt, William B StallcupAbstract:Cells expressing the NG2 Proteoglycan can attach, spread, and migrate on surfaces coated with NG2 mAbs, demonstrating that engagement of NG2 can trigger the cytoskeletal rearrangements necessary fo...
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a central segment of the NG2 Proteoglycan is critical for the ability of glioma cells to bind and migrate toward type vi collagen
Experimental Cell Research, 1997Co-Authors: Michael A Burg, Akiko Nishiyama, William B StallcupAbstract:Previous studies have established that the NG2 Proteoglycan binds directly to type VI collagen. To further our understanding of the biochemical and functional significance of this interaction we have used NG2 cDNA to construct a series of NG2 mutants with deletions spaced throughout the entire length of the 260-kDa NG2 core protein. Following transfection of these mutant cDNAs into B28 glioma cells, we determined the ability of mutant NG2 molecules to anchor type VI collagen on the cell surface. Eight of 11 transfectant populations were able to anchor type VI collagen. The three NG2 variants incapable of anchoring type VI collagen have deletions clustered within the central one-third of the NG2 ectodomain. These deletions identify a 469-amino-acid domain of NG2 responsible for binding of type VI collagen. Functional consequences of the NG2-type VI collagen interaction were explored by testing the relative ability of NG2-transfected and untransfected glioma cells to migrate toward type VI collagen. NG2-expressing cells exhibited a greater migratory response toward type VI collagen than their NG2-negative counterparts. This enhanced migration could be specifically inhibited with NG2 antibodies. Furthermore, glioma cells expressing NG2 in which the collagen-binding domain was deleted failed to exhibit this enhanced migration, whereas NG2 mutants in which non-collagen-binding regions were deleted continued to exhibit increased chemotaxis toward the type VI collagen. These comparisons confirm the importance of the central collagen-binding domain in mediating functionally important interactions between NG2 and type VI collagen.
Joel M. Levine - One of the best experts on this subject based on the ideXlab platform.
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entrapment via synaptic like connections between NG2 Proteoglycan cells and dystrophic axons in the lesion plays a role in regeneration failure after spinal cord injury
The Journal of Neuroscience, 2014Co-Authors: Angela R Filous, William B Stallcup, Amanda Phuong Tran, James C Howell, Sarah A Busch, Teresa A Evans, Shin H Kang, Dwight E Bergles, Seong Il Lee, Joel M. LevineAbstract:NG2 is purportedly one of the most growth-inhibitory chondroitin sulfate Proteoglycans (CSPGs) produced after spinal cord injury. Nonetheless, once the severed axon tips dieback from the lesion core into the penumbra they closely associate with NG2+ cells. We asked if Proteoglycans play a role in this tight cell—cell interaction and whether overadhesion upon these cells might participate in regeneration failure in rodents. Studies using varying ratios of CSPGs and adhesion molecules along with chondroitinase ABC, as well as purified adult cord-derived NG2 glia, demonstrate that CSPGs are involved in entrapping neurons. Once dystrophic axons become stabilized upon NG2+ cells, they form synaptic-like connections both in vitro and in vivo. In NG2 knock-out mice, sensory axons in the dorsal columns dieback further than their control counterparts. When axons are double conditioned to enhance their growth potential, some traverse the lesion core and express reduced amounts of synaptic proteins. Our studies suggest that Proteoglycan-mediated entrapment upon NG2+ cells is an additional obstacle to CNS axon regeneration.
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tpa mediated generation of plasmin is catalyzed by the Proteoglycan NG2
Glia, 2008Co-Authors: Westley B Nolin, Joel M. Levine, Jaime Emmetsberger, Noreen Bukhari, Yan Zhang, Stella E TsirkaAbstract:Paralysis resulting from spinal cord injury is devastating and persistent. One major reason for the inability of the body to heal this type of injury ensues from the local increase of glial cells leading to the formation of a glial scar, and the upregulation of chondroitin sulfate Proteoglycans (CSPGs) at the site of injury through which axons are unable to regenerate. Experimental approaches to overcome this problem have accordingly focused on reducing the inhibitory properties of CSPGs, for example by using chondroitinase to remove the sugar chains and reduce the CSPGs to their core protein constituents, although this step alone does not provide dramatic benefits as a monotherapy. Using in vitro and in vivo approaches, we describe here a potentially synergistic therapeutic opportunity based on tissue plasminogen activator (tPA), an extracellular protease that converts plasminogen (plg) into the active protease plasmin. We show that tPA and plg both bind to the CSPG protein NG2, which functions as a scaffold to accelerate the tPA-driven conversion of plg to plasmin. The binding occurs via the tPA and plg kringle domains to domain 2 of the NG2 CSPG core protein, and is enhanced in some settings after chondroitinase-mediated removal of the NG2 Proteoglycan side chains. Once generated, plasmin then degrades NG2, both in an in vitro setting using recombinant protein, and in vivo models of spinal cord injury. Our finding that the tPA and plg binding is in some instances more efficient after exposure of the NG2 Proteoglycan to chondroitinase treatment suggests that a combined therapeutic approach employing both chondroitinase and the tPA/plasmin proteolytic system could be of significant benefit in promoting axonal regeneration through glial scars after spinal cord injury.
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antibodies against the NG2 Proteoglycan promote the regeneration of sensory axons within the dorsal columns of the spinal cord
The Journal of Neuroscience, 2006Co-Authors: Andrew M Tan, Mario Colletti, Ann T Rorai, J Pate H Skene, Joel M. LevineAbstract:The NG2 chondroitin sulfate Proteoglycan inhibits axon growth in vitro. Levels of NG2 increase rapidly in the glial scars that form at sites of CNS injury, suggesting that NG2 may inhibit axon regeneration. To determine the functions of NG2, we infused mixtures of neutralizing or non-neutralizing anti-NG2 monoclonal antibodies into the dorsally transected adult rat spinal cord and analyzed the regeneration of ascending mechanosensory axons anatomically. At 1 week after injury, ascending sensory axons in control animals terminated caudal to the lesion within an area containing dense deposits of NG2 immunoreactivity. In animals treated with the neutralizing anti-NG2 antibodies, labeled axons penetrated the caudal border of the lesion and grew into and beyond the lesion center. The low intrinsic growth capacity of adult neurons may also limit the ability of damaged axons to regenerate. To enhance growth, we combined antibody treatment with a peripheral nerve conditioning lesion. After a conditioning lesion and treatment with control, non-neutralizing antibodies, many sensory axons grew into the lesion core. These axons did not grow past the rostral border of the lesion; rather, they grew along the dorsal surface of the spinal cord and within any remaining pieces of the dorsal roots. In contrast, combining a peripheral nerve conditioning lesion with neutralizing anti-NG2 antibodies resulted in sensory axon regeneration past the glial scar and into the white matter rostral to the injury site. The combinatorial approach used here that neutralizes extrinsic inhibition and increases intrinsic growth results in anatomically correct axon regeneration, a prerequisite for functional recovery.
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two separate metalloproteinase activities are responsible for the shedding and processing of the NG2 Proteoglycan in vitro
Molecular and Cellular Neuroscience, 2005Co-Authors: Richard A Asher, Joel M. Levine, Akiko Nishiyama, Daniel A Morgenstern, Francesca Properzi, James W FawcettAbstract:A high proportion of NG2 in the adult rat spinal cord is saline-soluble and migrates slightly faster than intact NG2 on SDS-PAGE, suggesting that it represents the shed ectodomain of NG2. In the injured cerebral cortex, much of the overall increase in NG2 is due to the saline-soluble (shed), rather than the detergent-soluble (intact), form. Hydroxamic acid metalloproteinase inhibitors, but not TIMPs, were able to prevent NG2 shedding in oligodendrocyte precursor cells (OPCs) in vitro. The generation of another truncated form of NG2 was, however, sensitive to TIMP-2 and TIMP-3. Two observations suggest that NG2 is involved in PDGF signaling in OPCs: the rate of NG2 shedding increased with cell density and NG2 expression was increased in the absence of PDGF. Ectodomain shedding converts NG2 into a diffusible entity able to interact with the growth cone, and we suggest that this release is likely to enhance its axon growth-inhibitory activity.
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expression and glycanation of the NG2 Proteoglycan in developing adult and damaged peripheral nerve
Molecular and Cellular Neuroscience, 2003Co-Authors: Daniel A Morgenstern, Joel M. Levine, Richard A Asher, Murali Naidu, Thomas Carlstedt, James W FawcettAbstract:We have investigated expression of the axon growth-inhibitory Proteoglycan NG2 in peripheral nerve. In the adult, NG2 was present on endoneurial and perineurial fibroblasts, but not on Schwann cells. At birth, peripheral nerve NG2 was heavily glycanated, but was much less so in the adult. In vitro, sciatic nerve fibroblasts also produced heavily glycanated NG2. After peripheral nerve injury in rats and humans, an accumulation of NG2-positive cells was observed at the injury site. In the rat, there was an increase in NG2 glycanation for at least 2 weeks following injury. In mixed cultures of Schwann cells and peripheral nerve fibroblasts, the axons preferred to grow on the Schwann cells and seldom crossed onto the fibroblasts. Three-dimensional cultures of sciatic nerve fibroblasts were inhibitory to the growth of dorsal root ganglion axons. Inhibition of Proteoglycan synthesis made the cells more permissive. NG2 may play a part in blocking axon regeneration through scar tissue in injured human peripheral nerve.
Michael A Burg - One of the best experts on this subject based on the ideXlab platform.
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cytoskeletal reorganization induced by engagement of the NG2 Proteoglycan leads to cell spreading and migration
Molecular Biology of the Cell, 1999Co-Authors: Xuexun Fang, Akiko Nishiyama, Kimberlee Dahlinhuppe, Michael A Burg, Diana Barritt, William B StallcupAbstract:Cells expressing the NG2 Proteoglycan can attach, spread, and migrate on surfaces coated with NG2 mAbs, demonstrating that engagement of NG2 can trigger the cytoskeletal rearrangements necessary fo...
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NG2 Proteoglycan binding peptides target tumor neovasculature
Cancer Research, 1999Co-Authors: Michael A Burg, Renata Pasqualini, Wadih Arap, Erkki Ruoslahti, William B StallcupAbstract:NG2 is the rat homologue of the human melanoma Proteoglycan, also known as the high molecular weight melanoma-associated antigen. This developmentally regulated membrane-spanning chondroitin sulfate Proteoglycan is expressed primarily by glial, muscle, and cartilage progenitor cells. Upon maturation, these cell types down-regulate NG2 expression. In adult animals, the expression of NG2 is restricted to tumor cells and angiogenic tumor vasculature, making this Proteoglycan a potential target for directing therapeutic agents to relevant sites of action. To this end, we have identified specific NG2-binding peptides by screening a phage-displayed random peptide library on purified NG2. Several rounds of biopanning on NG2 resulted in the specific enrichment of two phage-displayed decapeptides, TAASGVRSMH and LTLRWVGLMS. The binding of these phages to NG2 was inhibitable both by soluble NG2 and by glutathione S-transferase (GST) fusion proteins containing the cognate peptide sequences. In addition, direct binding between GST-TAASGVRSMH and GST-LTLRWVGLMS fusion proteins and NG2 was demonstrated in solid-phase binding assays. Interestingly, these NG2-binding fusion proteins cross-inhibited each other’s binding to NG2, suggesting that the two sequences bind to the same or overlapping sites on the Proteoglycan. Upon injection into tumor-bearing mice, NG2-binding phages specifically homed to tumor vasculature in wild-type mice but did not localize to the tumor vasculature in NG2 knockout mice. The in vivo targeting capability of these sequences suggests that they can be used for tumor targeting.
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high affinity binding of basic fibroblast growth factor and platelet derived growth factor aa to the core protein of the NG2 Proteoglycan
Journal of Biological Chemistry, 1999Co-Authors: Lothar Goretzki, Kathryn A Grako, Michael A Burg, William B StallcupAbstract:NG2 is a transmembrane chondroitin sulfate Proteoglycan that is expressed by immature progenitor cells in several developmental lineages and by some types of malignant cells. In vitro studies have suggested that NG2 participates in growth factor activation of the platelet-derived growth factor-α receptor. In this study the ability of recombinant NG2 core protein to interact with several different growth factors (epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor (PDGF)-AA, PDGF-BB, vascular endothelial growth factor (VEGF)165 and transforming growth factor (TGF)-β1) was investigated using two different assay systems: enzyme-linked immunosorbent assay-type solid-phase binding and an optical biosensor (BIAcore) system. High-affinity binding of bFGF and PDGF-AA to the core protein of NG2 could be demonstrated with both types of assays. Using both the BIAcore software analysis program and nonlinear regression analysis of the solid phase binding data, K D values in the low nanomolar range were obtained for binding of each of these growth factors to NG2. The results further indicate that NG2 contains at least two binding sites for each of these two growth factors. PDGF-BB, TGF-β1, VEGF, and EGF exhibited little or no binding to NG2 in either type of assay. These data suggest that NG2 can have an important role in organizing and presenting some types of mitogenic growth factors at the cell surface.
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expression of the NG2 Proteoglycan enhances the growth and metastatic properties of melanoma cells
Journal of Cellular Physiology, 1998Co-Authors: Michael A Burg, Kathryn A Grako, William B StallcupAbstract:The human homologue of NG2, the human melanoma Proteoglycan (HMP), is expressed on most human melanomas. To investigate the role of this Proteoglycan in melanoma progression, we have attempted to identify functionally important molecular ligands for NG2. Immunohistochemical analysis of cell lines that endogenously express NG2/HMP suggests that NG2/HMP associates with CD44 and α4β1 integrin, two molecules previously implicated in melanoma progression. Transfection of rat NG2 into the NG2-negative B16 mouse melanoma cell line also resulted in a highly colocalized pattern of expression between the transfected rat NG2 and the endogenously expressed mouse CD44 and α4β1 integrin molecules. In functional assays, expression of NG2 decreased the adhesion of B16 melanoma cells to CD44 monoclonal antibodies, hyaluronic acid, the C-terminal 40-kDa fibronectin fragment, and the CS1 fibronectin peptide, suggesting that NG2 may negatively modulate CD44- and α4β1-mediated binding events. Expression of NG2 increased the proliferation of melanoma cells in culture and increased tumorigenicity in vivo. Moreover, NG2 expression led to increased lung metastasis of B16F1 and B16F10 melanoma cells in experimental metastasis studies. Together, these studies demonstrate that NG2 is capable of modulating the adhesion, proliferation, and metastatic potential of melanoma cells. J. Cell. Physiol. 177:299–312, 1998. © 1998 Wiley-Liss, Inc.
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a central segment of the NG2 Proteoglycan is critical for the ability of glioma cells to bind and migrate toward type vi collagen
Experimental Cell Research, 1997Co-Authors: Michael A Burg, Akiko Nishiyama, William B StallcupAbstract:Previous studies have established that the NG2 Proteoglycan binds directly to type VI collagen. To further our understanding of the biochemical and functional significance of this interaction we have used NG2 cDNA to construct a series of NG2 mutants with deletions spaced throughout the entire length of the 260-kDa NG2 core protein. Following transfection of these mutant cDNAs into B28 glioma cells, we determined the ability of mutant NG2 molecules to anchor type VI collagen on the cell surface. Eight of 11 transfectant populations were able to anchor type VI collagen. The three NG2 variants incapable of anchoring type VI collagen have deletions clustered within the central one-third of the NG2 ectodomain. These deletions identify a 469-amino-acid domain of NG2 responsible for binding of type VI collagen. Functional consequences of the NG2-type VI collagen interaction were explored by testing the relative ability of NG2-transfected and untransfected glioma cells to migrate toward type VI collagen. NG2-expressing cells exhibited a greater migratory response toward type VI collagen than their NG2-negative counterparts. This enhanced migration could be specifically inhibited with NG2 antibodies. Furthermore, glioma cells expressing NG2 in which the collagen-binding domain was deleted failed to exhibit this enhanced migration, whereas NG2 mutants in which non-collagen-binding regions were deleted continued to exhibit increased chemotaxis toward the type VI collagen. These comparisons confirm the importance of the central collagen-binding domain in mediating functionally important interactions between NG2 and type VI collagen.
Krissa Gibby - One of the best experts on this subject based on the ideXlab platform.
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early vascular deficits are correlated with delayed mammary tumorigenesis in the mmtv pymt transgenic mouse following genetic ablation of the NG2 Proteoglycan
Breast Cancer Research, 2012Co-Authors: Weonkyoo You, Krissa Gibby, Kuniko Kadoya, Hildur Helgadottir, Lawrence Jt Young, Lesley G Ellies, Yunchao Chang, Robert D Cardiff, William B StallcupAbstract:Introduction The neuron-glial antigen 2 (NG2) Proteoglycan promotes pericyte recruitment and mediates pericyte interaction with endothelial cells. In the absence of NG2, blood vessel development is negatively impacted in several pathological models. Our goal in this study was to determine the effect of NG2 ablation on the early development and function of blood vessels in mammary tumors in the mammary tumor virus-driven polyoma middle T (MMTV-PyMT) transgenic mouse, and to correlate these vascular changes with alterations in mammary tumor growth.
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early vascular deficits are correlated with delayed mammary tumorigenesis in the mmtv pymt transgenic mouse following genetic ablation of the NG2 Proteoglycan
Breast Cancer Research, 2012Co-Authors: Weonkyoo You, Krissa Gibby, Kuniko Kadoya, Hildur Helgadottir, Lawrence Jt Young, Lesley G Ellies, Yunchao Chang, Robert D CardiffAbstract:The neuron-glial antigen 2 (NG2) Proteoglycan promotes pericyte recruitment and mediates pericyte interaction with endothelial cells. In the absence of NG2, blood vessel development is negatively impacted in several pathological models. Our goal in this study was to determine the effect of NG2 ablation on the early development and function of blood vessels in mammary tumors in the mammary tumor virus-driven polyoma middle T (MMTV-PyMT) transgenic mouse, and to correlate these vascular changes with alterations in mammary tumor growth. Three different tumor paradigms (spontaneous tumors, transplanted tumors, and orthotopic allografts of tumor cell lines) were used to investigate the effects of NG2 ablation on breast cancer progression in the MMTV-PyMT transgenic mouse. In addition to examining effects of NG2 ablation on mammary tumor growth, we also investigated effects on the structure and function of tumor vasculature. Ablation of NG2 led to reduced early progression of spontaneous, transplanted, and orthotopic allograft mammary tumors. NG2 was not expressed by the mammary tumor cells themselves, but instead was found on three components of the tumor stroma. Microvascular pericytes, myeloid cells, and adipocytes were NG2-positive in both mouse and human mammary tumor stroma. The effect of NG2 on tumor progression therefore must be stromal in nature. Ablation of NG2 had several negative effects on early development of the mammary tumor vasculature. In the absence of NG2, pericyte ensheathment of endothelial cells was reduced, along with reduced pericyte maturation, reduced sprouting of endothelial cells, reduced assembly of the vascular basal lamina, and reduced tumor vessel diameter. These early deficits in vessel structure are accompanied by increased vessel leakiness, increased tumor hypoxia, and decreased tumor growth. NG2 ablation also diminishes the number of tumor-associated and TEK tyrosine kinase endothelial (Tie2) expressing macrophages in mammary tumors, providing another possible mechanism for reducing tumor vascularization and growth. These results emphasize the importance of NG2 in mediating pericyte/endothelial cell communication that is required for proper vessel maturation and function. In the absence of normal pericyte/endothelial cell interaction, poor vascular function results in diminished early progression of mammary tumors.