Overexpression of Neurospora crassa OR74A glutamate decarboxylase in Escherichia coli for efficient GABA production

This study investigated the effect of glutamate decarboxylase from Neurospora crassa OR74A on GABA production in Escherichia coli. GABA is one of the inhibitory neurotransmitters in the mammalian central nervous system, and can be used as a precursor of promising biopolymer Nylon 4. E. coli that overexpressed N. crassa glutamate decarboxylase was cultured at various pH levels and temperatures to determine optimum conditions for GABA production. When the recombinant E. coli strain was cultured at 30°C and pH 3, a final GABA concentration of 5.26 g/L was obtained from 10 g/L of monosodium glutamate (MSG), corresponding to a GABA yield of 86.23%.     

Curcumin prevents liver fat accumulation and serum fetuin-A increase in rats fed a high-fat diet

Fetuin-A is synthesized in the liver and is secreted into the bloodstream. Clinical studies suggest involvement of fetuin-A in metabolic disorders such as visceral obesity, insulin resistance, diabetes, and fatty liver. Curcumin is extracted from the rhizome Curcuma longa and has been shown to possess potent antioxidant, anticarcinogenic, anti-inflammatory, and hypoglycemic properties. In this study, we investigated the effect of curcumin treatment on serum fetuin-A levels as well as hepatic lipids and prooxidant–antioxidant status in rats fed a high-fat diet (HFD). Male Sprague–Dawley rats were divided into six groups. Group 1 was fed control diet (10 % of total calories from fat). Groups 2 and 3 were given curcumin (100 and 400 mg/kg bw/day, respectively ) by gavage for 8 weeks and were fed control diet. Group 4 was fed with HFD (60 % of total calories from fat). Groups 5 and 6 received HFD together with the two doses of curcumin, respectively. Curcumin treatment appeared to be effective in reducing liver triglycerides and serum fetuin-A levels. These findings suggest that the reduction of fetuin-A may contribute to the beneficial effects of curcumin in the pathogenesis of obesity.     

A frequency monitoring system development for wide-area power grid protection

There have been recent research activities on GPS-based FNET to prevent wide-area blackouts by monitoring frequency deviation. This paper introduces a system for monitoring regional frequencies in power grid developed as an advanced research project for intelligent wide-area protective relaying in South Korea. The system is implemented by modeling an actual 345 kV transmission system of South Korea using EMTP-RV and by measuring voltages and currents at five regions. The frequencies are estimated using our previous proposed method, a frequency estimation algorithm with gain compensation. The monitoring system implemented is used to examine frequency propagation effects and various fault conditions. Simulation results showed that the proposed system could achieve good performance regarding the frequency monitoring under both steady states and dynamic fault conditions.     

High-yield production of the VP1 structural protein epitope from serotype O foot-and-mouth disease virus in Escherichia coli

For effective control of foot-and-mouth disease (FMD), the development of rapid diagnostic systems and vaccines are required against its etiological agent, FMD virus (FMDV). To accomplish this, efficient large-scale expression of the FMDV VP1 protein, with high solubility, needs to be optimized. We attempted to produce high levels of a serotype O FMDV VP1 epitope in Escherichia coli. We identified the subtype-independent serotype O FMDV VP1 epitope sequence and used it to construct a glutathione S-transferase (GST) fusion protein. For efficient production of the FMDV VP1 epitope fused to GST (VP1e–GST), four E. coli strains and three temperatures were examined. The conditions yielding the greatest level of VP1e–GST with highest solubility were achieved with E. coli BL21(DE3) at 25 °C. For high-level production, fed-batch cultures were conducted in 5-l bioreactors. When cells were induced at a high density and complex feeding solutions were supplied, approximately 11 g of VP1e–GST was obtained from a 2.9-l culture. Following purification, the VP1 epitope was used to immunize rabbits, and we confirmed that it induced an immune response.     

Plant species richness–productivity relationships in a low-productive boreal region

Local species richness–productivity (SR–P) relationship is usually reported as unimodal if long productivity gradients are sampled. However, it tends to be monotonically increasing in low-productive environments due to the decreasing part of the SR–P curve being truncated. Previous work indicated that this can hold true for forest herb layers, because of an upper bound on productivity caused mainly by canopy shading. Here, we ask whether the same pattern exists in a region with an upper bound on productivity caused by a harsh climate. We sampled herbaceous vegetation of boreal forests and grasslands in a low-productive region of central Yakutia (NE Siberia) with dry and winter-cool continental climate. We collected data on species composition, herb-layer productivity (aboveground herbaceous biomass), soil chemistry and light availability. We applied regression models to discriminate between monotonically increasing, decreasing and unimodal responses of herb-layer species richness to measured variables and analysed trends in the species-pool size and beta diversity along the productivity gradient. Our expectation of the monotonically increasing SR–P relationship was confirmed for neither forest herb layers nor grasslands. In the forest herb layers, no relationship was detected. In grasslands, the relationship was unimodal with species richness decline starting at much lower productivity levels than in more productive temperate grasslands. Potential causes for this decline are either limitation of local species richness by the species pool, which contains few species adapted to more productive habitats, or competitive exclusion, which can become an important control of species richness under lower levels of productivity than is the case in temperate grasslands.     

Circulating Matrix Metalloproteinase-2 and -9 Enzyme Activities in the Children with Ventricular Septal Defect

Ventricular septal defect (VSD) is the most common form of congenital heart diseases. Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases involved in causal cardiac tissue remodeling. We studied the changes of circulating MMP-2 and MMP-9 activities in the patients with VSD severity and closure. There were 96 children with perimembranous VSD enrolled in this study. We assigned the patients into three groups according to the ratio of VSD diameter/diameter of aortic root (Ao). They were classified as below: Trivial (VSD/Ao ratio ≤ 0.2), Small (0.2 < VSD/Ao ≤ 0.3) and Median (0.3 < VSD/Ao) group. Plasma MMP-2 and MMP-9 activities were assayed by gelatin zymography.There was a significant higher MMP-2 activity in the VSD (Trivial, Small and Median) groups compared with that in Control group. The plasma MMP-9 activity showed a similar trend as the findings in MMP-2 activity. After one year follow-up, a significant difference in the MMP-9 activity was found between VSD spontaneous closure and non-closure groups. In conclusion, a positive trend between the severity of VSD and activities of MMP-2 and MMP-9 was found. Our data imply that MMP-2 and MMP-9 activities may play a role in the pathogenesis of VSD.     

Oncogenes and inflammation rewire host energy metabolism in the tumor microenvironment

Here, we developed a model system to evaluate the metabolic effects of oncogene(s) on the host microenvironment. A matched set of “normal” and oncogenically transformed epithelial cell lines were co-cultured with human fibroblasts, to determine the “bystander” effects of oncogenes on stromal cells. ROS production and glucose uptake were measured by FACS analysis. In addition, expression of a panel of metabolic protein biomarkers (Caveolin-1, MCT1, and MCT4) was analyzed in parallel. Interestingly, oncogene activation in cancer cells was sufficient to induce the metabolic reprogramming of cancer-associated fibroblasts toward glycolysis, via oxidative stress. Evidence for “metabolic symbiosis” between oxidative cancer cells and glycolytic fibroblasts was provided by MCT1/4 immunostaining. As such, oncogenes drive the establishment of a stromal-epithelial “lactate-shuttle”, to fuel the anabolic growth of cancer cells. Similar results were obtained with two divergent oncogenes (RAS and NFκB), indicating that ROS production and inflammation metabolically converge on the tumor stroma, driving glycolysis and upregulation of MCT4. These findings make stromal MCT4 an attractive target for new drug discovery, as MCT4 is a shared endpoint for the metabolic effects of many oncogenic stimuli. Thus, diverse oncogenes stimulate a common metabolic response in the tumor stroma. Conversely, we also show that fibroblasts protect cancer cells against oncogenic stress and senescence by reducing ROS production in tumor cells. Ras-transformed cells were also able to metabolically reprogram normal adjacent epithelia, indicating that cancer cells can use either fibroblasts or epithelial cells as “partners” for metabolic symbiosis. The antioxidant N-acetyl-cysteine (NAC) selectively halted mitochondrial biogenesis in Ras-transformed cells, but not in normal epithelia. NAC also blocked stromal induction of MCT4, indicating that NAC effectively functions as an “MCT4 inhibitor”. Taken together, our data provide new strategies for achieving more effective anticancer therapy. We conclude that oncogenes enable cancer cells to behave as selfish “metabolic parasites”, like foreign organisms (bacteria, fungi, viruses). Thus, we should consider treating cancer like an infectious disease, with new classes of metabolically targeted “antibiotics” to selectively starve cancer cells. Our results provide new support for the “seed and soil” hypothesis, which was first proposed in 1889 by the English surgeon, Stephen Paget.     

Comparative Characteristics of Porous Bioceramics for an Osteogenic Response In Vitro and In Vivo

Porous calcium phosphate ceramics are used in orthopedic and craniofacial applications to treat bone loss, or in dental applications to replace missing teeth. The implantation of these materials, however, does not induce stem cell differentiation, so suitable additional materials such as porous calcium phosphate discs are needed to influence physicochemical responses or structural changes. Rabbit adipose-derived stem cells (ADSC) and mouse osteoblastic cells (MC3T3-E1) were evaluated in vitro by the MTT assay, semi-quantitative RT-PCR, and immunoblotting using cells cultured in medium supplemented with extracts from bioceramics, including calcium metaphosphate (CMP), hydroxyapatite (HA) and collagen-grafted HA (HA-col). In vivo evaluation of the bone forming capacity of these bioceramics in rat models using femur defects and intramuscular implants for 12 weeks was performed. Histological analysis showed that newly formed stromal-rich tissues were observed in all the implanted regions and that the implants showed positive immunoreaction against type I collagen and alkaline phosphatase (ALP). The intramuscular implant region, in particular, showed strong positive immunoreactivity for both type I collagen and ALP, which was further confirmed by mRNA expression and immunoblotting results, indicating that each bioceramic material enhanced osteogenesis stimulation. These results support our hypothesis that smart bioceramics can induce osteoconduction and osteoinduction in vivo, although mature bone formation, including lacunae, osteocytes, and mineralization, was not prominent until 12 weeks after implantation.