Perlecan domain V modulates astrogliosis In vitro and after focal cerebral ischemia through multiple receptors and increased nerve growth factor release

Astrogliosis constitutes part of the central nervous system's physiological response to injury. Considered for decades to be a major challenge for brain repair, recent studies have highlighted it as a promoter of such repair mechanisms. Recently, our group demonstrated the ability of perlecan domain V (DV) to be a novel potential stroke therapy by its neuroprotective effects. However, the potential for DV to modulate astrogliosis has not been investigated. The aim of this study is to better understand the relevance of DV to astrogliosis using both in vitro and in vivo rodent models. Notably, under basal conditions, astrocytes express all three DV receptors described in the literature: integrin α2β1, α5β1, and α‐dystroglycan (αDG). DV promoted astrocyte cell adhesion, cell migration as well as astrocyte stellation. Moreover, DV induced nerve growth factor (NGF) secretion through a αDG‐ and ERK‐dependent pathway. In contrast, α2β1 or α5β1 mediated DV antiproliferative effects in astrocytes. NGF production after DV treatment acted as a strong anti‐proliferative agent. Another remarkable effect of DV was that it decreased several markers of astrogliosis such as glial fibrillary acidic protein (GFAP), neurocan and phosphacan both in vitro and in vivo, suggesting the role of DV as a potential modulator of postinjury during late astrogliosis, and eventually the onset of glial scarring. Taken together, our study demonstrates the ability of DV to modulate key events of astrogliosis by promoting early astrogliosis and inhibiting glial scar formation, suggesting an additional therapeutic benefit of DV for recovery from stroke. © 2011 Wiley‐Liss, Inc.     

Immunohistochemical studies on proteoglycan expression in normal skin and chronic ulcers

BACKGROUND: Proteoglycans (PGs) represent a large family of complex molecules. They are found either as integral membrane components or constituents of the extracellular matrix. Their protein backbones are linked to different glycosaminoglycans, such as dermatan-, chondroitin-, keratan- or heparan sulphate. The molecules have specific functions during developmental processes as well as in diseases, such as cancer and inflammation. OBJECTIVES: The expression patterns of various cell-associated heparan and chondroitin/dermatan-sulphate PGs in human skin and chronic venous ulcers were investigated. METHODS: Tissue sections from 11 patients with chronic venous ulcers were used in this study. Monoclonal antibodies were used for detection of the proteoglycans syndecan-1, -2 and -4, glypican, CD44 and perlecan. RESULTS: The different PGs exhibited individual staining patterns. Syndecan-1 and -4 and glypican expression in chronic ulcers differed from the staining in normal skin. Whereas the expression of syndecan-4 and glypican in intact skin was mostly in the pericellular regions of keratinocytes, the epidermal cells from the wound edge contained mostly intracellular PGs. In the wound edge, syndecan-4 was predominantly expressed by epidermal basal layer cells. Syndecan-1 was less expressed at the epidermal wound margins. PGs bind growth factors, regulate proteolytic activity and act as matrix receptors. CONCLUSIONS: The altered expression patterns of glypican and syndecan-1 and -4 in chronic ulcers reflect their possible roles during inflammation and cell proliferation. Hence, analysis of PG expression should be of interest in future studies on normal as well as defective wound healing.     

串珠素在牙囊发育过程中的表达和意义

目的:观察牙囊发育不同时期串珠素(perlecan)分布、表达情况,探讨其与牙囊发育的关系。方法:出生0 d、1 d、1周、2周的SD大鼠取下颌第一磨牙及其牙囊组织,进行Perlecan免疫组化染色(SABC法),光镜观察其表达、分布,用图像分析测定其灰度值,并进行定量分析比较。结果:串珠素在各时间段牙囊均有阳性表达,而1 d、1周组表现为强阳性。图像分析结果表明1 d组、1周组大鼠阳性结果明显高于0 d组、2周组,出生后串珠素表达随着牙胚发育而不断增强,之后又随着牙胚的成熟而下降。结论:串珠素与牙囊的发育有关,在牙囊发育初期对牙囊基质的形成发挥作用,晚期随着基质的成熟其表达下降。     

The extracellular matrix proteoglycan perlecan facilitates transmembrane semaphorin-mediated repulsive guidance

The Drosophila transmembrane semaphorin-1a (Sema-1a) is a repulsive guidance cue that uses the Plexin A (PlexA) receptor during neural development. Sema-1a is required in axons to facilitate motor axon defasciculation at guidance choice points. We found that mutations in the trol gene strongly suppress Sema-1a-mediated repulsive axon guidance. trol encodes the phylogenetically conserved secreted heparan sulfate proteoglycan (HSPG) perlecan, a component of the extracellular matrix. Motor axon guidance defects in perlecan mutants resemble those observed in Sema-1a- and PlexA-null mutant embryos, and perlecan mutants genetically interact with PlexA and Sema-1a. Perlecan protein is found in both the CNS and the periphery, with higher expression levels in close proximity to motor axon trajectories and pathway choice points. Restoring perlecan to mutant motor neurons rescues perlecan axon guidance defects. Perlecan augments the reduction in phospho-focal adhesion kinase (phospho-FAK) levels that result from treating insect cells in vitro with Sema-1a, and genetic interactions among integrin, Sema-1a, and FAK in vivo support an antagonistic relationship between Sema-1a and integrin signaling. Therefore, perlecan is required for Sema-1a–PlexA-mediated repulsive guidance, revealing roles for extracellular matrix proteoglycans in modulating transmembrane guidance cue signaling during neural development.     

Spectrum of HSPG2 (Perlecan) mutations in patients with Schwartz-Jampel syndrome

Schwartz-Jampel syndrome (SJS) is a rare autosomal recessive condition defined by the association of myotonia with chondrodysplasia. SJS results from mutations in the HSPG2 gene, which encodes perlecan, a major component of basement membranes. Only eight HSPG2 mutations have been reported in six SJS families. Here, we describe the molecular findings in 23 families (35 patients) with SJS, being one-third of the SJS cases reported in the medical literature. We identified 22 new HSPG2 mutations and unreported polymorphisms. Mutations included nine deletion or insertion (41%), six splice site (27%), five missense (23%), and two nonsense mutations (9%). All but four mutations were private, and we found no evidence for a founder effect. Analyses of HSPG2 messenger RNA (mRNA) and perlecan immunostaining on patients' cells revealed a hypomorphic effect of the studied mutations. They also demonstrated distinct consequences of truncating and missense mutations on perlecan expression as truncating mutations resulted in instability of HSPG2 mRNA through nonsense mRNA-mediated decay, whereas missense mutations involving cysteine residues led to intracellular retention of perlecan, probably due to quality control pathways. Our analyses strengthen the idea that SJS results from hypomorphic mutations of the HSPG2 gene. They also propose tools for its molecular diagnosis and provide new clues for the understanding of its pathophysiology.     

Postnatal Development of the Lamina Reticularis in Primate Airways

The basement membrane zone (BMZ) appears as three component layers: the lamina lucida, lamina densa, and lamina reticularis. The laminas lucida and densa are present during all stages of development. The lamina reticularis appears during postnatal development. Collagens I, III, and V form heterogeneous fibers that account for the thickness of the lamina reticularis. Additionally, there are three proteoglycans considered as integral components of the BMZ: perlecan, collagen XVIII, and bamacan. Perlecan is the predominant heparan sulfate proteoglycan in the airway BMZ. It is responsible for many of the functions attributed to the BMZ, in particular, trafficking of growth factors and cytokines between epithelial and mesenchymal cells. Growth factor binding sites on perlecan include FGF‐1, FGF‐2, FGF‐7, FGF‐10, PDGF, HGF, HB‐EGF, VEGF, and TGF‐β. Growth factors pass through the BMZ when moving between the epithelial and mesenchymal cell layers. They move by rapid reversible binding with sites on both the heparan sulfate chains and core protein of perlecan. In this manner, perlecan regulates movement of growth factors between tissues. Another function of the BMZ is storage and regulation of FGF‐2. FGF‐2 has been shown to be involved with normal growth and thickening of the BMZ. Thickening of the BMZ is a feature of airway remodeling in asthma. It may have a positive effect by protecting against airway narrowing and air trapping. Conversely, it may have a negative effect by influencing trafficking of growth factors in the epithelial mesenchymal trophic unit. However, currently the significance of BMZ thickening is not known. Anat Rec, 293:947–954, 2008. © 2008 Wiley‐Liss, Inc.     

Perlecan and syndecan-4 in uterine tissues during the early pregnancy in mice

PROBLEM: During early pregnancy in mice, there is recruitment of specific immune cells, remodeling of the endometrium, cell differentiation and synthesis of new molecules. METHOD OF STUDY: Immunohistochemistry was used to determine the distribution of perlecan and syndecan-4 in the uteri before and after embryo implantation. RESULTS: During pre-implantation, perlecan was identified in basement membranes and extracellular spaces of the endometrial stroma. In contrast, expression of syndecan-4 was quite weak. In the peri-implantation period, perlecan remained in the basement membranes, and it was no longer observed in the stroma and it was identified in the embryonic cells. On day 4 of pregnancy, syndecan-4 increased in the fibroblasts of the subepithelial stroma. After implantation, syndecan-4 was pronounced in pre-decidual and mature decidual cells. CONCLUSIONS: The coordinate balance between the pre- and post-implantation periods suggests a role of these two molecules in the adaptive modification of the uterine microenvironment to receive and implant the embryo.     

Role of human airway smooth muscle in altered extracellular matrix production in asthma

1. The underlying abnormality in asthma is not fully understood; however, inflammation, airway remodelling and bronchial hyperresponsiveness are key factors. The plasma exudate from the microvascular leakage plays a significant role in remodelling, which includes extracellular matrix (ECM) protein deposition/breakdown and airway smooth muscle (ASM) hyperplasia/hypertrophy. 2. The ECM is an intricate network of macromolecules that forms the 'scaffolding' of the airways. This scaffolding not only acts as mechanical support that plays a crucial role in the maintenance of airway function and structure, but it is also a dynamic and complex network that has the potential to influence cellular function, including migration, differentiation and proliferation of a number of cell types. 3. In asthmatic airways, the profile of ECM proteins is altered. The deposition of collagen I, III, V, fibronectin, tenascin, hyaluronan, versican and laminin alpha2/beta2 is increased, whereas the deposition of collagen IV and elastin is decreased. 4. This imbalance in the ECM profile within the asthmatic airway could be due to: (i) increased de novo synthesis of ECM proteins; (ii) decreased activity of its degrading enzymes, namely matrix metalloproteinases (MMP); or (iii) upregulation of the tissue-specific inhibitors of metalloproteinases (TIMP). 5. One of the characteristic features of asthma is an increase in the amount of ASM within the airways. The ECM proteins/MMP/TIMP in and around the smooth muscle may play a contributory role in this increased growth. 6. The role of current asthma treatments in the prevention or reversal of airway ECM changes is an area that has only recently become of interest, with the majority of the in vivo work focusing on the effects of corticosteroids. 7. The evidence presented in this review indicates that the ASM may influence its own environment/proliferation through the production of ECM proteins, MMP and TIMP. Further studies are needed to fully understand the role of the ASM in the production of ECM proteins, MMP and TIMP andtheir potential influence in the mechanisms underlying asthma.