Nogo Receptor Inhibition Enhances Functional Recovery following Lysolecithin-Induced Demyelination in Mouse Optic Chiasm

Background

Inhibitory factors have been implicated in the failure of remyelination in demyelinating diseases. Myelin associated inhibitors act through a common receptor called Nogo receptor (NgR) that plays critical inhibitory roles in CNS plasticity. Here we investigated the effects of abrogating NgR inhibition in a non-immune model of focal demyelination in adult mouse optic chiasm.

Methodology/Principal Findings

A focal area of demyelination was induced in adult mouse optic chiasm by microinjection of lysolecithin. To knock down NgR levels, siRNAs against NgR were intracerebroventricularly administered via a permanent cannula over 14 days, Functional changes were monitored by electrophysiological recording of latency of visual evoked potentials (VEPs). Histological analysis was carried out 3, 7 and 14 days post demyelination lesion. To assess the effect of NgR inhibition on precursor cell repopulation, BrdU was administered to the animals prior to the demyelination induction. Inhibition of NgR significantly restored VEPs responses following optic chiasm demyelination. These findings were confirmed histologically by myelin specific staining. siNgR application resulted in a smaller lesion size compared to control. NgR inhibition significantly increased the numbers of BrdU+/Olig2+ progenitor cells in the lesioned area and in the neurogenic zone of the third ventricle. These progenitor cells (Olig2+ or GFAP+) migrated away from this area as a function of time.

Conclusions/Significance

Our results show that inhibition of NgR facilitate myelin repair in the demyelinated chiasm, with enhanced recruitment of proliferating cells to the lesion site. Thus, antagonizing NgR function could have therapeutic potential for demyelinating disorders such as Multiple Sclerosis.     

Asynchronous remodeling is a driver of failed regeneration in Duchenne muscular dystrophy

We sought to determine the mechanisms underlying failure of muscle regeneration that is observed in dystrophic muscle through hypothesis generation using muscle profiling data (human dystrophy and murine regeneration). We found that transforming growth factor β–centered networks strongly associated with pathological fibrosis and failed regeneration were also induced during normal regeneration but at distinct time points. We hypothesized that asynchronously regenerating microenvironments are an underlying driver of fibrosis and failed regeneration. We validated this hypothesis using an experimental model of focal asynchronous bouts of muscle regeneration in wild-type (WT) mice. A chronic inflammatory state and reduced mitochondrial oxidative capacity are observed in bouts separated by 4 d, whereas a chronic profibrotic state was seen in bouts separated by 10 d. Treatment of asynchronously remodeling WT muscle with either prednisone or VBP15 mitigated the molecular phenotype. Our asynchronous regeneration model for pathological fibrosis and muscle wasting in the muscular dystrophies is likely generalizable to tissue failure in chronic inflammatory states in other regenerative tissues.     

Evidence for a KATP Channel in Rough Endoplasmic Reticulum (rerKATP Channel) of Rat Hepatocytes

We report in a previous study the presence of a large conductance K⁺ channel in the membrane of rough endoplasmic reticulum (RER) from rat hepatocytes incorporated into lipid bilayers. Channel activity in this case was found to decrease in presence of ATP 100 µM on the cytoplasmic side and was totally inhibited at ATP concentrations greater than 0.25 mM. Although such features would be compatible with the presence of a K_ATP channel in the RER, recent data obtained from a brain mitochondrial inner membrane preparation have provided evidence for a Maxi-K channel which could also be blocked by ATP within the mM concentration range. A series of channel incorporation experiments was thus undertaken to determine if the ATP-sensitive channel originally observed in the RER corresponds to K_ATP channel. Our results indicate that the gating and permeation properties of this channel are unaffected by the addition of 800 nM charybdotoxin and 1 µM iberiotoxin, but appeared sensitive to 10 mM TEA and 2.5 mM ATP. Furthermore, adding 100 µM glibenclamide at positive potentials and 400 µM tolbutamide at negative or positive voltages caused a strong inhibition of channel activity. Finally Western blot analyses provided evidence for Kir6.2, SUR1 and/or SUR2B, and SUR2A expression in our RER fractions. It was concluded on the basis of these observations that the channel previously characterized in RER membranes corresponds to K_ATP, suggesting that opening of this channel may enhance Ca²⁺ releases, alter the dynamics of the Ca²⁺ transient and prevent accumulation of Ca²⁺ in the ER during Ca²⁺ overload.     

Stable expression of a fungal laccase protein using transplastomic tobacco

Laccase catalyzes the oxidation of various phenolic compounds that can be used in a wide range of industrial applications such as waste detoxification and the textile industry. In the present study, we generated transplastomic tobacco plants to develop a reliable commercial source of laccase production. The stability of the laccase protein in the transgenic plants was increased by using the enhancer sequence from green fluorescent protein, resulting in three independent lines with high levels of laccase accumulation (up to 2?% of total protein); significant laccase activity, however, was not detected. Interestingly, the transplastomic lines showed slightly retarded vegetative growth, with a light green leaf color in comparison with the control, which may be attributable to copper deficiency induced by ligand chelation by abundantly produced laccase. These results suggest that the tobacco chloroplast is an efficient system for the mass production of laccase protein, but further studies are needed to obtain active enzyme.     

Genetic and Chemical Diversity in Perovskia abrotanoides Kar. (Lamiaceae) Populations Based on ISSRs Markers and Essential Oils Profile

Genetic and the essential oil composition variability among twelve Perovskia abrotanoides populations (PAbPs) growing wild in Iran were assessed by ISSR markers, GC‐FID and GC/MS, respectively. Nine selected ISSR primers produced 119 discernible bands, of them 96 (80.7%) being polymorphic. Genetic similarity values among populations ranged between 0.07 and 0.79 which indicated a high level of genetic variation. Polymorphic information content, resolving power and marker index generated by ISSR primers were, 0.31, 6.14, and 3.32, respectively. UPGMA grouped PAbPs into four main clusters. Altogether, 38 chemical compounds were identified in the oils, and a relatively high variation in their contents was found. Camphor (11.9 – 27.5%), 1,8‐cineole (11.3 – 21.3%), α‐bisabolol (0.0 – 13.1%), α‐pinene (5.9 – 10.8%), and δ‐3‐carene (0.1 – 10.5%) were the major compounds. Oxygenated monoterpenes (32.1 – 35.8%) and monoterpene hydrocarbons (25.7 – 30.4%) were the main groups of compounds in the oils studied. Cluster analysis and principal‐component analysis were used to characterize the samples according to oil components. Four main chemotypes were found to be Chemotype I (camphor/1,8‐cineol), Chemotype II (1,8‐cineole/camphor), Chemotype III (camphor/1,8‐cineol/α‐bisabolol), and Chemotype IV (camphor/δ‐3‐carene/α‐bisabolol). The information, provided here on P. abrotanoides populations, will be useful to introduce this plant into agricultural systems.     

Effects of sodium benzoate,a commonly used food preservative,on learning,memory, and oxidative stress in brain of mice

Sodium benzoate (SB) is a widely used preservative and antimicrobial substance in many foods and soft drinks. However, this compound is generally recognized as safe food additives, but evidence has suggested that a high intake of SB may link to attention deficit‐hyperactivity disorder in children. Present study investigate the effects of oral administration of different concentrations of SB (0.56, 1.125, and 2.25 mg/mL) for 4 weeks, on the learning and memory performance tests, and also the levels of malondialdehyde (MDA), reduced glutathione (GSH), and acetylcholinesterase activity (AChE) in the mouse brain. The results showed that SB significantly impaired memory and motor coordination. Moreover, SB decreased reduced GSH and increased the MDA level in the brain significantly (P < 0.001). However, nonsignificant alteration was observed in the AChE activity. These findings suggest that short‐term consumption of SB can impair memory performance and increased brain oxidative stress in mice.     

Down-regulation of 14-3-3 zeta sensitizes human glioblastoma cells to apoptosis induction

Strong 14-3-3 zeta protein expression plays an important role in tumorigenesis, including in the maintenance of cell growth, resistance increase, and the prevention of apoptosis. In this study, we focus on two targets: (1) the expression of 14-3-3 zeta in the different grades of human astrocytoma (II–IV), (2) suppression of 14-3-3 zeta protein expression in glioblastoma derived astrocytes by 14-3-3 zeta shRNA lentiviral particles. The tissues of human astrocytoma were provided from 30 patients (ten of each grade of astrocytoma). Control tissues were obtained from the peritumoral brain zone of those patients with glioblastoma. The protein and mRNA expression levels of each astrocytoma grade were assessed via western blotting and RT-PCR, respectively. Results indicated that 14-3-3 zeta was significantly expressed in glioblastoma multiforme (grade IV) and 14-3-3 zeta expression levels enhanced according to the increase of astrocytoma malignancy. In the cellular study for knock down of the 14-3-3 zeta protein, surgical biopsy of glioblastoma was used to isolate primary astrocyte. Astrocytes were transduced with 14-3-3 zeta shRNA or non-targeted shRNA lentiviral particles. Furthermore, reduction of the 14-3-3 zeta protein expression in the astrocytes evaluated through qRT-PCR and western blot after transduction of 14-3-3 zeta shRNA lentiviral particles. Moreover, apoptosis properties, including DNA fragmentation and ratio increase of Bax/Bcl-2 were observed in astrocytes following reduction of 14-3-3 zeta protein expression. Further observation indicated that the mitochondrial pathway through release of cytochorome c and caspase-3 activity was involved in the apoptosis induction. Hence, this study demonstrates a key role of the 14-3-3 zeta protein in tumorigenesis but also indicates that 14-3-3 zeta can be considered as a target for the astrocytoma treatment specially glioblastoma.     

Optimization of protein extraction from Gelidiella acerosa by carbohydrases using response surface methodology

In this study, application of response surface methodology for enzymic pretreatment optimization of Gelidiella acerosa was investigated in order to improve the extraction of algal proteins using Viscozyme L and Celluclast 1.5L. The total protein, soluble proteins and reducing sugar recovery in the water‐soluble fraction were studied in relation to the hydrolysis time, type and concentration of the enzymes. Enzymatic digestion appeared to be an effective treatment for protein extraction. While enzyme hydrolysis by Celluclast 1.5L was able to facilitate the protein extraction, it was a relatively inefficient way to improve protein extraction yield, in comparison with Viscozyme L. The optimum conditions for protein extraction was found to be hydrolysis by 2.8 μL mL^?1 of Viscozyme L for 12 h.     

Graphene oxide/silver nanohybrid: Optimization,antibacterial activity and its impregnation on bacterial cellulose as a potential wound dressing based on GO‐Ag nanocomposite‐coated BC

Recently, bacterial cellulose (BC) based wound dressing have raised significant interests in medical fields. However, to our best knowledge, it is apparent that the BC itself has no antibacterial activity. In this study, we optimized graphene oxide‐silver (GO‐Ag) nanohybrid synthesis using Response Surface Methodology and impregnate it to BC and carefully investigate their antibacterial activities against both the Gram‐negative bacteria Escherichia coli and the Gram‐positive bacteria Staphylococcus aureus. We discover that, compared to silver nanoparticles, GO‐Ag nanohybrid with an optimal GO suspension's pH and ratio is much more effective and shows synergistically enhanced, strong antibacterial activities at rather low dose. The GO‐Ag nanohybrid is more toxic to E. coli than that to S. aureus. The antibacterial and mechanical properties of BC/GO‐Ag composite are further investigated.