Gender dimorphism influences extracellular matrix expression and regeneration of muscular tissue in mdx dystrophic mice

Mdx mouse, the animal model of Duchenne muscular dystrophy, lacks dystrophin and develops an X-linked recessive inflammatory myopathy characterized by degeneration of skeletal muscle fibers and connective tissue replacement. The present work aimed to assess whether gender dimorphism in mdx mice would influence skeletal muscle pathology at ages corresponding to main histological changes in the microenvironment of muscular tissue: myonecrosis, regeneration, and fibrosis. At the height of myonecrosis (6 weeks postnatal), skeletal muscles of male mdx mice showed increased sarcolemmal permeability, numerous inflammatory foci, and marked deposition of the extracellular matrix components (ECM) type I collagen and laminin. In contrast, age-matched mdx females showed mild ECM deposition, discrete myonecrosis, but increased numbers of regenerating fibers expressing the satellite cell marker NCAM. In contrast ovariectomized mdx females showed decreased numbers of regenerating fibers. Older (24 and 48 weeks postnatal) mdx females showed extensive fibrosis with increased sarcolemmal permeability and marked deposition of ECM components than corresponding males. These results suggest a role for female hormones in the control of myonecrosis probably by promoting regeneration of muscular tissue and mitigating inflammation especially at ages under the critical influence of sex hormones.     

提高转基因表达策略研究进展

随着转基因技术在各个应用领域的深入和发展 ,转基因沉默现象已引起越来越多人的关注。在介绍转基因沉默的概念、分子机理的基础上 ,着重对提高转基因表达的方法、策略及其研究进展进行综述。分别阐述内含子、组织特异性启动子、增强子、定点整合、DNA大片段、反式作用因子以及基因组调控元件核基质附着区对转基因表达的影响。为克服转基因失活、提高转基因的表达效率提供理论依据。     

Oxidative and Nitrosative Modifications of Tropoelastin Prevent Elastic Fiber Assembly in Vitro

Elastic fibers are extracellular structures that provide stretch and recoil properties of tissues, such as lungs, arteries, and skin. Elastin is the predominant component of elastic fibers. Tropoelastin (TE), the precursor of elastin, is synthesized mainly during late fetal and early postnatal stages. The turnover of elastin in normal adult tissues is minimal. However, in several pathological conditions often associated with inflammation and oxidative stress, elastogenesis is re-initiated, but newly synthesized elastic fibers appear abnormal. We sought to determine the effects of reactive oxygen and nitrogen species (ROS/RNS) on the assembly of TE into elastic fibers. Immunoblot analyses showed that TE is oxidatively and nitrosatively modified by peroxynitrite (ONOO⁻) and hypochlorous acid (HOCl) and by activated monocytes and macrophages via release of ONOO⁻ and HOCl. In an in vitro elastic fiber assembly model, oxidatively modified TE was unable to form elastic fibers. Oxidation of TE enhanced coacervation, an early step in elastic fiber assembly, but reduced cross-linking and interactions with other proteins required for elastic fiber assembly, including fibulin-4, fibulin-5, and fibrillin-2. These findings establish that ROS/RNS can modify TE and that these modifications affect the assembly of elastic fibers. Thus, we speculate that oxidative stress may contribute to the abnormal structure and function of elastic fibers in pathological conditions.     

Durable and broad-spectrum disease protection measure against airborne phytopathogenic fungi by using the detachment action of gelatinolytic bacteria

We had previously obtained collagenolytic/gelatinolytic bacteria, which degrade the fungal extracellular matrix, to establish a novel biological control measure that inhibits germling adhesion of airborne phytopathogenic fungi on the host plant surface. By using barley-Magnaporthe oryzae pathosystem, Chryseobacterium sp. was most effective biocontrol agents as tested. The selected bacteria were evaluated for durable disease protection against M. oryzae on barley leaves by using chloramphenicol-resistant mutants. Chryseobacterium sp. from the soil was less likely to settle on leaf surfaces. Therefore, we tried to manipulate Chryseobacterium sp. to inhabit the leaf’s surface. The gelatin supplementation dramatically improved the settlement of gelatinolytic bacteria Chryseobacterium sp. from the soil, and the disease protection effect lasted for more than 2 weeks on barley. Moreover, exploitation of Chryseobacterium sp. for disease protection was extended against other airborne pathogens, Alternaria alternata Japanese pear pathotype on Japanese pear and Colletotrichum orbiculare on cucumber.     

Developmental and Age-Related Changes in Rat Brain Glycosaminoglycans

The quantities of each major class of glycosaminoglycan were determined in rat cerebrum from postnatal day 5 to 30 months of age. Chondroitin sulphate, dermatan sulphate, heparan sulphate, heparin, and hyaluronate were found, but no keratan sulphate was detected. Large and rapid changes in glycosaminoglycan content were observed during the period of brain maturation, and thereafter relatively steady levels were maintained until after the age of 12 months. The most remarkable change in the aged rat cerebrum was the ratio by weight of hyaluronate to chondroitin sulphate, which was approximately 1:1 from postnatal day 10 to 18 months but increased to 2.6:1 by the age of 30 months. In immature rats, the proportion of nonsulphated and 6-sulphated disaccharides derived from chondroitinase AC digests of brain glycosaminoglycans was much greater than in adults. In mature rats, chondroitin sulphate was composed almost entirely of 4-sulphated disaccharide subunits. The possibility that these changes could affect the permeability properties of the cerebral extracellular space and ionic equilibria in the brain is discussed.     

Differential expression of extracellular-signal-regulated kinase 5 (ERK5) in normal and degenerated human nucleus pulposus tissues and cells

Extracellular-signal-regulated kinase 5 (ERK5) is a member of the mitogen-activated protein kinase (MAPK) family and regulates a wide variety of cellular processes such as proliferation, differentiation, necrosis, apoptosis and degeneration. However, the expression of ERK5 and its role in degenerated human nucleus pulposus (NP) is hitherto unknown. In this study, we observed the differential expression of ERK5 in normal and degenerated human nucleus pulposus tissues by using immunohistochemical staining and Western blot. Treatment of NP cells with Pro-inflammatory cytokine, TNF-α decreased ERK5 gene expression as well as NP marker gene expression; including the type II collagen and aggrecan. Suppression of ERK5 gene expression in NP cells by ERK5 siRNA resulted in decreased gene expression of type II collagen and aggrecan. Furthermore, inhibition of ERK5 activation by BIX02188 (5 μM) decreased the gene expression of type II collagen and aggrecan in NP cells. Our results document the expression of ERK5 in degenerated nucleus pulposus tissues, and suggest a potential involvement of ERK5 in human degenerated nucleus pulposus.     

Interleukin-6 (IL-6) Trans Signaling Drives a STAT3-dependent Pathway That Leads to Hyperactive Transforming Growth Factor-β (TGF-β) Signaling Promoting SMAD3 Activation and Fibrosis via Gremlin Protein

Fibrosis is a common and intractable condition associated with various pathologies. It is characterized by accumulation of an excessive amount of extracellular matrix molecules that primarily include collagen type I. IL-6 is a profibrotic cytokine that is elevated in the prototypic fibrotic autoimmune condition systemic sclerosis and is known to induce collagen I expression, but the mechanism(s) behind this induction are currently unknown. Using healthy dermal fibroblasts in vitro, we analyzed the signaling pathways that underscore the IL-6-mediated induction of collagen. We show that IL-6 trans signaling is important and that the effect is dependent on STAT3; however, the effect is indirect and mediated through enhanced TGF-β signaling and the classic downstream cellular mediator Smad3. This is due to induction of the bone morphogenetic protein (BMP) antagonist Gremlin-1, and we show that Gremlin-1 is profibrotic and is mediated through canonical TGF-β signaling.     

Enhanced assay of endothelial exocytosis using extracellular matrix components

Vascular inflammation plays a key role in the pathogenesis of atherosclerosis. The first step in vascular inflammation is endothelial exocytosis, in which endothelial granules fuse with the plasma membrane, releasing prothrombotic and proinflammatory messenger molecules. The development of cell culture models to study endothelial exocytosis has been challenging because the factors that modulate exocytosis in vitro are not well understood. Here we report a method for studying endothelial exocytosis that optimizes extracellular matrix components, cell density, and duration of culture. Human umbilical vein endothelial cells plated on collagen I-coated plates and cultured in the confluent state for 7–12 days in low-serum medium showed robust secretion of von Willebrand factor when stimulated with various agonists. This exocytosis assay is rapid and applicable to high-throughput screening.     

Cell adhesion: old and new questions

Metazoans clearly need cell adhesion to hold themselves together, but adhesion does much more than that. Adhesion receptors make transmembrane connections, linking extracellular matrix and adjacent cells to the intracellular cytoskeleton, and they also serve as signal transducers. In this article, I briefly summarize our present understanding of the molecular basis and biological consequences of cell adhesion and discuss how our current knowledge sheds light on questions of specificity of cell adhesion. I offer some thoughts and speculations about the evolution of cell-adhesion molecules and processes, consider their inter-relationships with other forms of cell–cell communication and discuss unresolved questions ripe for investigation as we enter the postgenomic era.