The multifaceted role of astrocytes in regulating myelination

Astrocytes are the major glial cell of the central nervous system (CNS), providing both metabolic and physical support to other neural cells. After injury, astrocytes become reactive and express a continuum of phenotypes which may be supportive or inhibitory to CNS repair. This review will focus on the ability of astrocytes to influence myelination in the context of specific secreted factors, cytokines and other neural cell targets within the CNS. In particular, we focus on how astrocytes provide energy and cholesterol to neurons, influence synaptogenesis, affect oligodendrocyte biology and instigate cross-talk between the many cellular components of the CNS

Astroglial-axonal interactions during early stages of myelination in mixed cultures using in vitro and ex vivo imaging techniques

<b>Background</b><p></p> Myelination is a very complex process that requires the cross talk between various neural cell types. Previously, using cytosolic or membrane associated GFP tagged neurospheres, we followed the interaction of oligodendrocytes with axons using time-lapse imaging in vitro and ex vivo and demonstrated dynamic changes in cell morphology. In this study we focus on GFP tagged astrocytes differentiated from neurospheres and their interactions with axons.<p></p> <b>Results</b><p></p> We show the close interaction of astrocyte processes with axons and with oligodendrocytes in mixed mouse spinal cord cultures with formation of membrane blebs as previously seen for oligodendrocytes in the same cultures. When GFP-tagged neurospheres were transplanted into the spinal cord of the dysmyelinated shiverer mouse, confirmation of dynamic changes in cell morphology was provided and a prevalence for astrocyte differentiation compared with oligodendroglial differentiation around the injection site. Furthermore, we were able to image GFP tagged neural cells in vivo after transplantation and the cells exhibited similar membrane changes as cells visualised in vitro and ex vivo.<p></p> <b>Conclusion</b><p></p> These data show that astrocytes exhibit dynamic cell process movement and changes in their membrane topography as they interact with axons and oligodendrocytes during the process of myelination, with the first demonstration of bleb formation in astrocytes

Subcellular localization of Mayven following expression of wild type and mutant EGFP tagged cDNAs

<p>Abstract</p> <p>Background</p> <p>Process formation by glial cells is crucial to their function. Mayven, an actin binding, multi-domain polypeptide, and member of the BTB-BACK-Kelch family have been shown to be important in oligodendrocyte process extension. To assess the role of Mayven in neural cell process extension we have tracked the subcellular distribution of exogenous Mayven following expression of a rat Mayven -EGFP cDNA in a variety of neural cell backgrounds and specifically in OEC tranfectants following drug treatment to disrupt the integrity of the cytoskeleton. A comparison was made between the subcellular localization following transient transfection of OECs with full-length Mayven cDNA and a series of mutant domain constructs.</p> <p>Results</p> <p>The subcellular location of Mayven in OEC transfectants showed a characteristic distribution with intense foci of staining towards the process tips corresponding to regions of accumulated Mayven overlapping in part with lammelipodial actin and was absent from the filipodia and the outer membrane. This signature pattern was also observed in Schwann cells, Oli-Neu cells, astrocytes and the neuroblastoma cell line B104 transfectants and resembled the exogenous and endogenous Mayven distribution in oligodendrocytes. This contrasted with the localization pattern in non-neural cells. There was a re-localization of Mayven in OEC transfectants following drug treatment to challenge the integrity of the actin cytoskeleton while breakdown of the microtubular component had no discernible impact on the accumulation of Mayven in the process tips. Deletion of the first three amino acids of the SH3 motif of the putative Fyn Kinase binding domain at the amino terminus significantly compromised this signature pattern as did the removal of the last Kelch repeat unit of six unit Kelch domain comprising the carboxyl terminus. In addition, there was a reduction in process length in mutant transfectants. Co-expression studies with a haemagglutinin (HA) tagged wild type Mayven cDNA and EGFP tagged mutant cDNAs suggested a homomeric interaction mediated by the BTB/POZ domain.</p> <p>Conclusions</p> <p>Exogenous Mayven is transported to the lamellipodia in neural transfectants associating with the actin cytoskeletal network. In addition to the importance of the internal BTB/POZ domain, this subcellular distribution pattern is dependent on the presence of an intact amino and carboxyl terminus.</p

The therapeutic potential of niche-specific mesenchymal stromal cells for spinal cord injury repair

The use of mesenchymal stem/stromal cells (MSCs) for transplant-mediated repair represents an important and promising therapeutic strategy after spinal cord injury (SCI). The appeal of MSCs has been fuelled by their ease of isolation, immunosuppressive properties, and low immunogenicity, alongside the large variety of available tissue sources. However, despite reported similarities in vitro, MSCs sourced from distinct tissues may not have comparable biological properties in vivo. There is accumulating evidence that stemness, plasticity, immunogenicity, and adaptability of stem cells is largely controlled by tissue niche. The extrinsic impact of cellular niche for MSC repair potential is therefore important, not least because of its impact on ex vivo expansion for therapeutic purposes. It is likely certain niche-targeted MSCs are more suited for SCI transplant-mediated repair due to their intrinsic capabilities, such as inherent neurogenic properties. In addition, the various MSC anatomical locations means that differences in harvest and culture procedures can make cross-comparison of pre-clinical data difficult. Since a clinical grade MSC product is inextricably linked with its manufacture, it is imperative that cells can be made relatively easily using appropriate materials. We discuss these issues and highlight the importance of identifying the appropriate niche-specific MSC type for SCI repair

Calponin is expressed by subpopulations of connective tissue cells but not olfactory ensheathing cells in the neonatal olfactory mucosa

&lt;b&gt;Background&lt;/b&gt;&lt;br /&gt; Debate has been ongoing on the relative merits of olfactory ensheathing cells (OECs) and Schwann cells as candidates for transplant-mediate repair of CNS lesions. Both glial cells exhibit similar molecular and cellular properties and to date there has been no antigenic marker identified that can clearly distinguish the two cell types. This inability to distinguish between the two cells types prevents confirmation of a controversial statement that cultures of OECs are contaminated with Schwann cells. Recently, proteomic analysis of foetal OECs and adult Schwann cells identified an actin-binding protein, calponin, as a specific marker for OECs. However, at the same time a recent report suggested that adult OECs do not express calponin. It was not clear if this discrepancy was due to methodology, as cells had to be treated with proteinase K to maximize calponin staining or developmental differences with only foetal/neonatal OECs expressing calponin. For this reason we have examined calponin expression in the peripheral olfactory system of embryonic and neonatal rats &lt;i&gt;in vivo&lt;/i&gt; and from cells &lt;i&gt;in vitro&lt;/i&gt; to assess if calponin is expressed in a developmental manner.&lt;p&gt;&lt;/p&gt; &lt;b&gt;Results&lt;/b&gt;&lt;br /&gt; In this study we show that: i) proteinase K pretreatment had no effect on calponin staining in both OECs and Schwann cells. ii) calponin immunoreactivity was not expressed by embryonic or neonatal OECs &lt;i&gt;in vitro&lt;/i&gt; and &lt;i&gt;in vivo&lt;/i&gt; although connective tissue from the olfactory mucosa was strongly positive in neonatal rats but not embryonic rats, iii) calponin expression in the olfactory mucosa was heterogeneous, defining subpopulations of connective tissue cells iv) using functional confrontation assays between OECs or Schwann cells with astrocytes, calponin was expressed heterogeneously by astrocytes.&lt;p&gt;&lt;/p&gt; &lt;b&gt;Conclusion&lt;/b&gt;&lt;br /&gt; It is concluded that calponin is heterogeneously expressed by neonatal mucosal connective tissue but not expressed by neonatal OECs, embryonic OECs, and neonatal Schwann cells. Furthermore, we propose that calponin is not a specific marker for OECs generated from any developmental age.&lt;p&gt;&lt;/p&gt

Role of IL-33 and ST2 signalling pathway in multiple sclerosis: expression by oligodendrocytes and inhibition of myelination in central nervous system

Recent research findings have provided convincing evidence indicating a role for Interleukin-33 (IL-33) signalling pathway in a number of central nervous system (CNS) diseases including multiple sclerosis (MS) and Alzheimer’s disease. However, the exact function of IL-33 molecule within the CNS under normal and pathological conditions is currently unknown. In this study, we have mapped cellular expression of IL-33 and its receptor ST2 by immunohistochemistry in the brain tissues of MS patients and appropriate controls; and investigated the functional significance of these findings in vitro using a myelinating culture system. Our results demonstrate that IL-33 is expressed by neurons, astrocytes and microglia as well as oligodendrocytes, while ST2 is expressed in the lesions by oligodendrocytes and within and around axons. Furthermore, the expression levels and patterns of IL-33 and ST2 in the lesions of acute and chronic MS patient brain samples are enhanced compared with the healthy brain tissues. Finally, our data using rat myelinating co-cultures suggest that IL-33 may play an important role in MS development by inhibiting CNS myelination

Review of Dam Effects on Native and Invasive Crayfishes Illustrates Complex Choices for Conservation Planning

Dams are among the most prevalent and extreme alterations humans have perpetrated on fluvial systems. The dramatic physical and biological changes caused by dams have been synthesized for many aquatic faunal groups, but not for crayfishes. In addition, invasive crayfish species are an increasing threat to global biodiversity, and dams have both costs and benefits with respect to crayfish invasions. North American crayfishes have imperiled native crayfishes in Europe, largely by hosting and spreading the crayfish plague pathogen Aphanomyces astaci that is lethal to European crayfishes. The differential effects of A. astaci on North American vs. European crayfishes contribute to differences between the continents in the costs and benefits of dams. We reviewed literature on both the detrimental and beneficial effects of dams on crayfishes, with emphasis on conservation of European crayfishes. We also suggested additional potential dam effects that warrant investigation. Our review illustrates the challenges and opportunities dams create for crayfish conservation. Dams create detrimental effects to native crayfishes, including reducing suitable habitats necessary for native habitat-specialist species and creating habitats suitable for non-native habitat-generalist species; fragmenting crayfish populations; and reducing species' ability to recolonize upstream habitats. Conversely, dams can have beneficial effects by creating barriers that slow or halt upstream invasions by non-native crayfishes and spread of the crayfish plague. The complexity of the issues and the limited ecological information available highlights the need for future studies on the effects of dams on crayfishes. Crayfishes are one of the most imperiled groups of aquatic fauna globally; therefore, understanding the beneficial and detrimental effects of dams is essential for effective conservation of many crayfish species

The development of a ε-polycaprolactone (PCL) scaffold for CNS repair

Potential treatment strategies for the repair of spinal cord injury (SCI) currently favour a combinatorial approach incorporating several factors, including exogenous cell transplantation and biocompatible scaffolds. The use of scaffolds for bridging the gap at the injury site is very appealing although there has been little investigation into CNS neural cell interaction and survival on such scaffolds before implantation. Previously we demonstrated that aligned micro-grooves 12.5-25 µm wide on ε-polycaprolactone (PCL) promoted aligned neurite orientation and supported myelination. In this study we identify the appropriate substrate and its topographical features required for the design of a 3D scaffold intended for transplantation in SCI. Using an established myelinating culture system of dissociated spinal cord cells, recapitulating many of the features of the intact spinal cord, we demonstrate that astrocytes plated on the topography secrete soluble factors(s) that delay oligodendrocyte differentiation but do not prevent myelination. However, as myelination does occur after a further 10-12 days in culture this does not prevent the use of PCL as a scaffold material as part of a combined strategy for the repair of SCI

The use of myelinating cultures as a screen of glycomolecules for CNS repair

In vitro cell-based assays have been fundamental in modern drug discovery and have led to the identification of novel therapeutics. We have developed complex mixed central nervous system (CNS) cultures, which recapitulate the normal process of myelination over time and allow the study of several parameters associated with CNS damage, both during development and after injury or disease. In particular, they have been used as a reliable screen to identify drug candidates that may promote (re)myelination and/or neurite outgrowth. Previously, using these cultures, we demonstrated that a panel of low sulphated heparin mimetics, with structures similar to heparan sulphates (HSs), can reduce astrogliosis, and promote myelination and neurite outgrowth. HSs reside in either the extracellular matrix or on the surface of cells and are thought to modulate cell signaling by both sequestering ligands, and acting as co-factors in the formation of ligand-receptor complexes. In this study, we have used these cultures as a screen to address the repair potential of numerous other commercially available sulphated glycomolecules, namely heparosans, ulvans, and fucoidans. These compounds are all known to have certain characteristics that mimic cellular glycosaminoglycans, similar to heparin mimetics. We show that the N-sulphated heparosans promoted myelination. However, O-sulphated heparosans did not affect myelination but promoted neurite outgrowth, indicating the importance of structure in HS function. Moreover, neither highly sulphated ulvans nor fucoidans had any effect on remyelination but CX-01, a low sulphated porcine intestinal heparin, promoted remyelination in vitro. These data illustrate the use of myelinating cultures as a screen and demonstrate the potential of heparin mimetics as CNS therapeutics

Species Tropism of Chimeric SHIV Clones Containing HIV-1 Subtype-A and Subtype-E Envelope Genes

AbstractTo analyze HIV-1 genes in a nonhuman primate model for lentivirus infection and AIDS, recombinant SIV/HIV-1 (SHIV) clones were constructed from two HIV-1 subtype-A isolates (HIV-1SF170 and HIV-1Q23–17 from individuals in Africa) and two HIV-1 subtype-E isolates (HIV-19466 and HIV-1CAR402 from AIDS patients in Thailand and Africa), respectively. These four SHIV clones, designated SHIV-A-170, SHIV-A-Q23, SHIV-9466.33, and SHIV-E-CAR, contain envelope (env) genes from the subtype-A or -E viruses. Interestingly, SHIV-A-170, SHIV-A-Q23, and SHIV-9466.33 were restricted for replication in cultures of macaque lymphoid cells, whereas SHIV-E-CAR replicated efficiently in these cells. Additional studies to define the block to replication in macaque cells were focused on the subtype-E clone SHIV-9466.33. A SHIV intragenic env clone, containing sequence-encompassing V1/V2 regions of HIV-1CAR402 and V3/V4/V5 regions of SHIV-9466.33, infected and replicated in macaque lymphoid cells. These results indicated that the sequence-encompassing V1/V2 region of HIV-19466 was responsible for the block of the SHIV-9466.33 replication in macaque cells. Analysis of viral DNA in acutely infected macaque cells revealed that SHIV-9466.33 was blocked at a step at/or before viral DNA synthesis, presumably during the process of virion entry into cells. In a fluorescence-based cell–cell fusion assay, fusion pore formation readily took place in cocultures of cells expressing the SHIV-9466.33 env glycoprotein with macaque T-lymphoid cells. Taken together, these results demonstrated that the block of SHIV-9466.33 replication in macaque cells is at an early step after fusion pore formation but before reverse transcription