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A stereodivergent Pd/Cu catalyst system for asymmetric desymmetric alkylation of allylic geminal dicarboxylates has been developed, which was successfully applied to the asymmetric synthesis of β-h...

3,3-Disubstituted oxindoles bearing quaternary and tertiary stereogenic centers are privileged structural motifs, which widely exist in pharmaceutical and natural products. Herein, a highly regio-, enantio-, and diastereoselective allylic alkylation of 3-alkyl oxindoles through synergistic iridium and copper catalysis is described, which provides a series of 3,3-disubstituted oxindole derivatives containing adjacent quaternary and tertiary stereogenic centers in excellent yields, enantiomeric excess, and diastereomeric ratio (for 30 examples, up to 97 % yield, >99 % ee, and >20 : 1 dr). This method provides exclusive branched selectivity, excellent enantio- and diastereoselectivities, and good functional compatibility. Control experiments suggested that the chiral copper catalyst is required for achieving high reactivities and diastereoselectivities under mild reaction conditions.

Preeclampsia (PE) is a serious pregnancy-related disease, and patients usually present with a high inflammatory response. Previous studies have suggested that aspirin (ASP) may have a role in alleviating the pathogenesis of preeclampsia. However, whether ASP can improve kidney damage and the mechanism for improving it is currently unclear. Here we optimized a lipopolysaccharide (LPS)-induced PE mouse model to identify the role of ASP in renal protection. We found that ASP treatment ameliorated LPS-induced renal failure and pathological changes, the tubular injury was significantly attenuated by ASP. Administration of ASP decreased the renal expression of pro-inflammatory factors, resulting in reduced kidney inflammation. The number of GALECTIN-3-positive cells was reduced, and the up-regulation of IL-6 and TNF-α was decreased. In addition, ASP also suppressed renal cell apoptosis and oxidative stress. An in vitro study indicated that ASP relieved LPS-induced HK-2 cell damage by inhibiting WNT5A/NF-κB signaling. Collectively, our data suggest that ASP is a useful therapeutic option for PE-related kidney injury.

We report a stereodivergent Pd/Cu catalyst system that was successfully applied to the asymmetric allylic alkylation of symmetrical 1,3-disubstituted allyl acetates with prochiral imino esters, providing efficient access to enantiopure products bearing vicinal stereocenters in a fully stereodivergent manner. The protocol proceeds smoothly under mild reaction conditions and can accommodate a range of imino esters, delivering the substituted products in high yields and with excellent diastereoselectivities (up to >20 : 1 dr) and enantioselectivities (up to >99% ee).

This study aimed to comprehensively understand the proteomic characteristics and modulation of the neural microenvironment with N-methyl- d -aspartate (NMDA)-induced neuronal degeneration in the retina and optic nerve at 12 h after intravitreal injection of 40 nmol NMDA. Male Sprague–Dawley rats were sacrificed at 12 h after intravitreal injection of 40 nmol NMDA. PBS-injected eyes served as controls. The key cell death-linked proteins from the retina and optic nerve tissues were assessed by a mass spectrometry-based label-free approach. In proteomics analysis, we identified 3532 proteins in retinal tissues and 2593 proteins in optic nerve tissues. The ACSL3 (Q63151) and Prnp (P13852) proteins were upregulated in the NMDA-damaged retina and connected with ferroptosis. The Gabarapl2 (P60522) protein was upregulated in NMDA-damaged optic nerves and connected with autophagy. We performed parallel reaction monitoring (PRM) to validate the liquid chromatography-tandem mass spectrometry (LC–MS/MS) results. Data are available ProteomeXchange with identifiers PXD022466 (label-free quantification) and PXD022729 (PRM validation). Significance Excitotoxicity is one of the pathogeneses of various retinal disorders, including glaucoma, retinal ischemia-reperfusion and traumatic optic neuropathy. This study indicated that ferroptosis may be linked to pathological cell death in the retina with NMDA insult. Autophagy may be induced by NMDA overstimulation in both the optic nerve and retina. Regulating these types of death simultaneously may provide the maximum benefit for retinal disease therapy.

Esterases are a large family of enzymes with wide applications in the industry. However, all esterases originated from natural sources, limiting their use in harsh environments or newly- emerged reactions. In this study, we designed a new esterase to develop a new protocol to satisfy the needs for better biocatalysts. The ideal spatial conformation of the serine catalytic triad and the oxygen anion hole at the substrate-binding site was constructed by quantum mechanical calculation. The catalytic triad and oxygen anion holes were then embedded in the protein scaffold using the new enzyme protocol in Rosetta 3. The design results were subsequently evaluated, and optimized designs were used for expression and purification. The designed esterase had significant lytic activities towards p-nitrophenyl acetate, which was confirmed by point mutations. Thus, this study developed a new protocol to obtain novel enzymes that may be useful in unforgiving environments or novel reactions.

Hydrofunctionalisation and difunctionalisation of dienes, allenes, and alkynes are widely utilized in the synthesis of valuable allylic compounds. In the past decades, asymmetric catalysis has emerged as one of the most attractive fields in organic synthesis. Recently, the asymmetric versions of hydrofunctionalisation and difunctionalisation reactions have become powerful and compelling tools to afford enantiopure allylic compounds, appealing to a large range of chemists. Various metal complexes modified with a large number of chiral ligands and several chiral organocatalysts have been developed to promote the hydrofunctionalisation and difunctionalisation reactions and expand substrate scope. This review provides an overview of this field, and aims at summarizing the chiral ligand used in this area of research. A detailed discussion of the development of these reactions and the general reaction mechanisms is provided.

The double-layer membrane consisting of sulfonated poly(ether sulfone) (SPES) sub-layer and polyvinyl alcohol (PVA) sub-layer (denoted as SPES/PVA membrane) was prepared and employed as the separator for vanadium redox flow battery (VRB) system to evaluate the vanadium ions permeability and cell performance. The SPES/PVA membrane is a double-layer structure and exhibits dramatically lower vanadium ions permeability and better cell performance compared to the pristine SPES membrane, PVA membrane, and Nafion117 membrane. The vanadium ion permeability of SPES/PVA membrane is one order of magnitude lower than that of Nafion117 membrane. In further work, the single cell with SPES/PVA membrane showed significantly lower capacity loss, higher coulombic efficiency (>92.5%), and higher energy efficiency (>83.9%) than Nafion117 membrane. In the self-discharge test, SPES/PVA membrane showed 1.8 times longer duration in the open circuit decay than Nafion117 membrane. With all the good properties and low cost, this new kind of double-layer membrane is suggested to have excellent commercial prospects as an ion exchange membrane for VRB systems.

Lipase is one of the most widely used classes of enzymes in biotechnological applications and organic chemistry. Candida rugosa lipases (CRL) can catalyze hydrolysis, esterification and transesterification with high regio-, stereo- and enantio-selectivity. However, thermal inactivation above 45 °C limits CRL's applications. Studies on improving the thermal stability of CRL are often limited by its slow-growing eukaryotic expression host, which is not suitable for large-scale screening. Identification of thermally stable mutants by rational design, regarded as an efficient substitution of experimental efforts, would provide a method for site-directed improvement of CRL. In this study, mutation-induced stability changes in CRL Lip1 were predicted by three rational design methods. Followed by conservative analyses and functional region exclusion, five mutants of a hot-spot, Asp457Phe, Asp457Trp, Asp457Met, Asp457Leu, and Asp457Tyr, were identified and prepared for enzymatic characterization. These five mutants increased the apparent melting temperature of Lip1 from 7.4 °C to 9.3 °C, with the most thermostable mutant, Asp457Phe, exhibiting a 5.5-fold longer half-life at 50 °C and a 10 °C increase in optimum temperature. Furthermore, pH stability of Lip1 was also enhanced due to the introduction of Asp457Phe mutation. The study demonstrates that thermally stable mutants of CRL could be identified with limited experimental efforts using rational design methods.

Soil microorganisms are major drivers of soil carbon (C) cycling; however, the response of these microorganisms to climate change remains unclear. In the present study, we investigated how 18 months of multifactor climate treatments (warmed air temperature by 3 °C and decreased or increased precipitation manipulation by 30%) affected soil microbial biomass C and nitrogen (N), community substrate utilization patterns, and community composition. Decreased and increased precipitation significantly reduced microbial biomass C by 13.5% and 24.9% and microbial biomass N by 22.9% and 17.6% in unwarmed plots, respectively (P < 0.01). Warming enhanced community substrate utilization by 89.8%, 20.4%, and 141.4% in the natural, decreased, and increased precipitation plots, respectively. Particularly, warming significantly enhanced the utilization of amine and carboxylic acid substrates among all precipitation manipulation plots. Compared with the natural air temperature with natural precipitation treatment, other treatments affected fungal community richness by −0.9% to 33.6% and reduced the relative abundance of the dominant bacterial and fungal groups by 0.5% to 6.8% and 4.3% to 10.7%, respectively. The warming and/or precipitation manipulation treatments significantly altered Zygomycota abundance (P < 0.05). Our results indicate that climate change drivers and their interactions may cause changes in soil microbial biomass C and N, community substrate utilization patterns, and community composition, particularly for the fungal community, and shifts in the microorganism community may further shape the ecosystems function. Scopus

Most natural proteins exhibit poor thermostability, which limits their industrial application. Computer-aided rational design is an efficient purpose-oriented method that can improve protein thermostability. Numerous machine-learning-based methods have been designed to predict the changes in protein thermostability induced by mutations. However, all of these methods have certain limitations due to existing mutation coding methods that overlook protein sequence features. Here we propose a method to predict protein thermostability using convolutional neural networks based on an in-depth study of thermostability-related protein properties. This method comprises a three-dimensional coding algorithm, including protein mutation information and a strategy to extract neighboring features at protein mutation sites based on multiscale convolution. The accuracies on the S1615 and S388 data sets, which are widely used for protein thermostability predictions, reached 86.4 and 87%, respectively. The Matthews correlation coefficient was nearly double those produced using other methods. Furthermore, a model was constructed to predict the thermostability of Rhizomucor miehei lipase mutants based on the S3661 data set, a single amino acid mutation data set screened from the ProTherm protein thermodynamics database. Compared with the RIF strategy, which consists of three algorithms, i.e., Rosetta ddg monomer, I Mutant 3.0, and FoldX, the accuracy of the proposed method was higher (75.0 vs 66.7%), and the negative sample resolution was simultaneously enhanced. These results indicate that our prediction method more effectively assessed the protein thermostability and distinguished its features, making it a powerful tool to devise mutations that enhance the thermostability of proteins, particularly enzymes.

In this study, biodiesel production from Chinese tallow kernel oil catalyzed by Candida rugosa lipase (CRL) in ionic liquid was investigated. Biodiesel was prepared from Chinese tallow kernel oil. Nineteen kinds of ionic liquids were screened for CRL to enhance its catalysis ability for biodiesel synthesis. Interestingly, [Hmim][PF6], not only help to simplify the downstream processing, but also boost the efficiency of CRL from about 35% to 95.4%. In addition, the biodiesel mixed with various blending ratios of fossil diesel was used to fuel the conventional diesel engine to examine fuel and emission properties. Most of fuel characteristics of the biodiesel met the standards set by EN and ASTM. Its output power was found to be close to pure diesel. The emission products, including hydrocarbons (HC, 52.7% lower) and carbon monoxide (CO, 48.6% lower), decreased substantially with the exception of oxynitride (NOx) emission, which displayed a 12.8% increase. The results suggest that biodiesel production from woody oil catalyzed by CRL in [Hmim][PF6], is feasible in future application with a promising prospect.

Pre-eclampsia (PE) is a serious hypertensive disorder of pregnancy that remains a leading cause of perinatal and maternal morbidity and mortality worldwide. Placental ischemia/hypoxia and the secretion of soluble fms-like tyrosine kinase 1 (sFlt1) into maternal circulation are involved in the pathogenesis of PE. Although low-dose aspirin (LDA) has beneficial effects on the prevention of PE, the exact mechanisms of action of LDA, particularly on placental dysfunction, and sFlt1 release, have not been well investigated. This study aimed to determine whether LDA exists the protective effects on placental trophoblast and endothelial functions and prevents PE-associated sFlt1 release. First, we observed that LDA mitigated hypoxia-induced trophoblast apoptosis, showed positive effects on trophoblast cells migration and invasion activity, and increased the tube-forming activity of human umbilical vein endothelial cells (HUVECs). In addition, LDA decreased hypoxia-induced sFlt1 production, and the c-Jun NH2 -terminal kinase/activator protein-1 (JNK/AP-1) pathway was shown to mediate the induction of sFlt1. Moreover, the transcription factor AP-1 was confirmed to regulate the Flt1 gene expression by directly binding to the Flt1 promoter in luciferase assays. The result of chromatin immunoprecipitation assays further demonstrated that LDA could directly decrease the expression of the transcription factor AP-1, and thus decrease sFlt1 production. Finally, the effects of LDA on sFlt1 production were proved in human placental explants. Taken together, our data show the protective effects of LDA against trophoblast and endothelial cell dysfunction and reveal that the LDA-mediated inhibition of sFlt1 via the JNK/AP-1 pathway may be a potential cellular/molecular mechanism for the prevention of PE.

Gestational diabetes mellitus (GDM) is the primary cause of maternal and fetal morbidity and mortality. Insulin resistance (IR) is pivotal to GDM pathogenesis, and mammalian target of rapamycin (mTOR) is a critical regulator of GDM. An increasing amount of evidence indicates that vitamin D deficiency is a risk factor for GDM. However, there are few reports on the effect of IR on GDM placentas. The present study aims to verify that 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) can ameliorate trophoblast IR by suppressing mTOR signaling. An IR BeWo cell model was established in the presence of high insulin and glucose medium. The IR level and mTOR activation with or without 1,25(OH)2D3 treatment were evaluated. The IR cells exhibited excessive mTOR signaling activation, upregulated inflammatory factor levels, and extensive lipid infiltration. However, 1,25(OH)2D3 reversed mTOR activation and reduced the IR level and lipid infiltration. In addition, 1,25(OH)2D3 treatment in GDM placental explants blocked the aberrant, increased levels of leptin, TNF-α, and IL-6. Therefore, 1,25(OH)2D3 treatment protects trophoblasts against high IR mainly through suppressing mTOR signaling, and this mechanism may serve as a potential therapy for patients with GDM.

International audience; AbstractKey messageThinning increased microbial biomass but did not alter enzyme activities in the soil ofPinus densifloraSieb. et Zucc. forests in South Korea. This effect of thinning was larger under a relatively heavy thinning intensity, but there was divergence in the magnitude between sites.ContextThe balance between microbial biomass accumulation and enzymatic C and N assimilation determines the level of bio-available C and N. However, the effects of thinning on these parameters remain contradictory and unconfirmed.AimsThe effects of thinning intensity on microbial biomass and enzyme activity were assessed in the soil of Pinus densiflora Sieb. et Zucc. forests in South Korea.MethodsUn-thinned control and 15 and 30% basal area thinning treatments were applied to two 51- to 60-year-old P. densiflora forests with different management histories, topographies, rainfall amounts, and soils. Seven years after thinning, microbial biomass and activities of N-acetyl-glucosaminidase, β-glucosidase, cellobiohydrolase, β-xylosidase, phenol oxidase, and peroxidase were measured before and after seasonally concentrated rains and at 0–10 cm depth.ResultsMicrobial biomass was generally highest under the 30% basal area thinning and lowest under the control, and was positively correlated to total soil C and N. The increase in microbial biomass was lower at the site displaying sandier, drier, and more acidic soils and retaining smaller amounts of thinning residue. Conversely, thinning had no significant effect on activities of all enzymes at both sites in both periods.ConclusionThinning can promote accumulation of microbial biomass without significant change in enzyme activities participating in the assimilation of C and N. This effect of thinning tended to increase with thinning intensity but differed in magnitude between sites.

Air warming (TC: control; TW: +3 °C) and precipitation manipulation (PC: control; PD: −30%; PI: +30%) were established to examine effects of these treatments on fine root production (FRP), fine root mortality (FRM), and total root (coarse and fine root) biomass in 33- to 59-month-old Pinus densiflora Sieb. et Zucc. seedlings for two years. We hypothesized that warming and altered precipitation would affect the growth, death, and biomass of fine roots by changing soil temperature and soil water availability. Mean annual FRP and total root biomass were significantly altered by only precipitation manipulation: they were 29.3% (during the two-year period) and 69.0% (after the entire two years) higher, respectively, in PD plots than in PC plots, respectively. In contrast, only warming had a significant effect on mean annual FRM, being 13.2% lower in TW plots than TC plots during the two-year period. Meanwhile, fine root biomass was affected negatively and simultaneously by both soil temperature and soil moisture. It seemed that fine root dynamics have changed so that they maintain their systems in response to the altered soil temperature and moisture. The current study adds significant knowledge for understanding the fine root dynamics of P. densiflora seedlings under altered temperature and precipitation regimes.

Diseases associated with Abelson-related gene (also called ABL2) include leukemia; furthermore, previous researches have studied the expressions and functions of ABL2 in different types of malignancies and found that it plays an important role in almost all kinds of cancers. Nevertheless, the mechanism of ABL2 in gastric cancer (GC) remains vague. In the present study, the level of ABL2 in human GC tissues was detected by immunohistochemistry. Also, the GC cell lines MGC-803 and BGC-823 were selected to stably knock down and overexpress the level of ABL2 by corresponding lentiviral vectors. Puromycin was used to maintain the stable low expression of ABL2 MGC-803 cells compared with control cells; what is more, the high expression of ABL2 BGC-823 cells was also obtained. Based on it, we detected the proteins associated with apoptosis, such as Bcl-2 family and caspase family by western blotting. The most appropriate concentration of puromycin to kill GC cells is 1 µg/mL; then, we obtained the corresponding stable cell lines. Furthermore, we found that high level of ABL2 in BGC-823 cells increased the expression of Bcl-XL, total PARP, and caspase3, while decreased the level of cleaved caspase3 and cleaved caspase9. Consistent results are received in MGC-803 cells. In addition, ABL2 overexpression led to the protein related with Ras/Erk and PI3K/AKT signaling pathway increased; also, we found that the major proteins play a significant role in it. All the data showed that high expression of ABL2 suppresses apoptosis through Ras/Erk and PI3K/AKT signaling pathway in GC cell lines.

Industrialization of enzyme-catalyzed biodiesel production has been hindered by low conversion efficiency and long reaction time. As a representative example, Rhizopus oryzae lipase (ROL) exhibited a poor performance in biodiesel production. However, higher yields and shorter reaction time could be achieved by screening different lipases that could synergically catalyze the reaction with ROL. The results showed that Novozym 435 combined with ROL displayed the best performance, with a biodiesel yield that was 30% higher than that of ROL alone in 30 h. Hence, the use of a synergic strategy was investigated systematically. Under optimized conditions, the biodiesel yield was as high as 98.3% and the reaction time was successfully shortened from 60 h to 21 h. Moreover, the combined lipases retained a yield of about 80% after 20 cycles in a solvent-free system. In addition to ROL, the performance of other lipases whose regioselectivity were similar to ROL was improved when they were combined with Novozym 435. In conclusion, synergic catalysis is a promising strategy for enhancing biodiesel yield and minimizing reaction time for ROL.

Rhizomucor miehei lipase (RML), as a kind of eukaryotic protein catalyst, plays an important role in the food, organic chemical, and biofuel industries. However, RML retains its catalytic activity below 50°C, which limits its industrial applications at higher temperatures. Soluble expression of this eukaryotic protein in Escherichia coli not only helps to screen for thermostable mutants quickly but also provides the opportunity to develop rapid and effective ways to enhance the thermal stability of eukaryotic proteins. Therefore, in this study, RML was engineered using multiple computational design methods, followed by filtration via conservation analysis and functional region assessment. We successfully obtained a limited screening library (only 36 candidates) to validate thermostable single point mutants, among which 24 of the candidates showed higher thermostability and 13 point mutations resulted in an apparent melting temperature ( T m app ) of at least 1°C higher. Furthermore, both of the two disulfide bonds predicted from four rational-design algorithms were further introduced and found to stabilize RML. The most stable mutant, with T18K/T22I/E230I/S56C-N63C/V189C-D238C mutations, exhibited a 14.3°C-higher T m app and a 12.5-fold increase in half-life at 70°C. The catalytic efficiency of the engineered lipase was 39% higher than that of the wild type. The results demonstrate that rationally designed point mutations and disulfide bonds can effectively reduce the number of screened clones to enhance the thermostability of RML. IMPORTANCE R. miehei lipase, whose structure is well established, can be widely applied in diverse chemical processes. Soluble expression of R. miehei lipase in E. coli provides an opportunity to explore efficient methods for enhancing eukaryotic protein thermostability. This study highlights a strategy that combines computational algorithms to predict single point mutations and disulfide bonds in RML without losing catalytic activity. Through this strategy, an RML variant with greatly enhanced thermostability was obtained. This study provides a competitive alternative for wild-type RML in practical applications and further a rapid and effective strategy for thermostability engineering.

Although coarse woody debris (CWD) is important for soil functioning, the mechanism which affects soil properties beneath CWD are unclear. Here, initial changes in microbial and soil properties were studied using homogenous CWD samples in eight Korean red pine (Pinus densiflora Sieb. et Zucc.) forests. For each forest, CWD samples (diameter: 11.1 ± 0.1 cm; length: 10.2 ± 0.0 cm) from similarly aged Korean red pine trees were laid on the mineral soil surface from May to June, 2016, and soils were sampled at points beneath CWD and at a distance of 1 m from the CWD after 1 year. Soils beneath the CWD had higher moisture but lower inorganic nitrogen (N) and a higher microbial biomass C (carbon)/N ratio than those sampled 1 m from the CWD. No differences in total C and N, labile C, pH, and C substrate utilization between the soils were significant. The difference in inorganic N between the soils decreased with increasing CWD decomposition, whereas that for microbial biomass fraction in total C and N increased correspondingly. Our results showed that soil microbial affinity for retaining N might become higher than that for retaining C under the presence of CWD, which possibly alters N availability and generates a spatial heterogeneity in forest soils.

In this study, hydrophobic macroporous resin NKA was employed as matrix for immobilization of free Rhizopus oryzae lipase (ROL). The performance of the immobilized ROL was significantly enhanced. The recovery activity was up to 1,293.78 % and the specific activity increased to 152,914 U/g-protein, which was 46-fold higher than that of the free lipase. Moreover, the immobilized lipase showed higher thermostability and better pH-resistance than its free counterpart. Additionally, three different nonaqueous modification strategies (including bioimprinting, lecithin coating, and lyophilization protection) were further utilized to improve the performance of the immobilized lipase. The corresponding enhancements were 33.68 %, 31.98 %, and 99.86 %. When these modifications were combined together, the activity improved 209.51 %. In order to confirm its practical application, the modified ROL was used to biorefine rapeseed oil deodorizer distillate (RODD) for biodiesel production. The highest conversion yield reached 98.23 %, much close to that (97.46 %) of Novozym 435. The results suggest that the prepared lipase in this study is a promising biocatalyst with high stability, efficiency and operational reusability.

To examine the effect of exopolysaccharides (EPS) on the microstructure and textural and rheological properties of yoghurt, purified EPS were added to buffalo milk to make yoghurts. The results showed that the viscosity, water-holding capacity, hardness, and microstructure of yoghurt were distinctly affected by the level of addition of EPS. With the increasing amount of supplemented EPS, the viscosity of buffalo yoghurt showed a downward trend with the increasing shear velocity, and an increasing degree of recovery of viscosity with the decrease in shear velocity, indicating that the steric barrier formed between the EPS and protein enhanced the viscoelastic properties. The microstructure showed more dense interspaced voids with EPS addition.

As the decomposition of lignocellulosic compounds is a rate-limiting stage in the nutrient mineralization from organic matters, elucidation of the changes in soil enzyme activity can provide insight into the nutrient dynamics and ecosystem functioning. The current study aimed to assess the effect of thinning intensities on soil conditions. Un-thinned control, 20 % thinning, and 30 % thinning treatments were applied to a Larix kaempferi forest, and total carbon and nitrogen, total carbon to total nitrogen ratio, extractable nutrients (inorganic nitrogen, phosphorus, calcium, magnesium, potassium), and enzyme activities (acid phosphatase, β-glucosidase, β-xylosidase, β-glucosaminidase) were investigated. Total carbon and nitrogen concentrations were significantly increased in the 30 % thinning treatment, whereas both the 20 and 30 % thinning treatments did not change total carbon to total nitrogen ratio. Inorganic nitrogen and extractable calcium and magnesium concentrations were significantly increased in the 20 % thinning treatment; however, no significant changes were found for extractable phosphorus and potassium concentrations either in the 20 or the 30 % thinning treatment. However, the applied thinning intensities had no significant influences on acid phosphatase, β-glucosidase, β-xylosidase, and β-glucosaminidase activities. These results indicated that thinning can elevate soil organic matter quantity and nutrient availability, and different thinning intensities may affect extractable soil nutrients inconsistently. The results also demonstrated that such inconsistent patterns in extractable nutrient concentrations after thinning might not be fully explained by the shifts in the enzyme-mediated nutrient mineralization.

The aim of this study was to investigate the short-term effects of warming treatment and precipitation manipulation on the growth, photosynthetic rate, and chlorophyll content of Pinus densiflora seedlings based on a climate change scenario in Korea. Two year old P. densiflora seedlings were planted in a nursery in April 2013. The air temperature of warmed plots (W) was set to increase by 3 °C compared to control plots (C) using an infrared heater in May 2013. The three precipitation manipulations consisted of decreased precipitation using transparent panels (–30%; P–), increased precipitation using pumps and drip irrigation (+30%; P+), and a control (0%; P0). Root collar diameter and seedling height of P. densiflora were measured in April and October 2013. Net photosynthetic rate and total chlorophyll contents were measured from June to October 2013. Warming treatment increased the growth of root collar diameter, and the interaction effect of the warming treatment and precipitation manipulation on it was also significant. In contrast, no significant effects were found for the growth of seedling height. These results demonstrate that warming treatment might affect the growth of root collar diameter and seedling height differently. It was also found that WP– treatment decreased net photosynthetic rates. This pattern might be due to the soil moisture availability, as soil moisture content was lowest in the WP– treatment and net photosynthetic rate was elevated with increasing soil moisture content. Meanwhile, warming treatment increased the total chlorophyll content. The results suggest that chlorophyll synthesis was promoted by a temperature increase in the leaves. The current study, which simulated a climate change environment in an open field, provides important information for the prediction of the ecophysiological effects of changes in temperature and precipitation on P. densiflora seedlings in Korea. This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) that is funded by the Ministry of Education (2013R1A1A2012242) and by the Korea Forest Service (S111115L030100). Scopus

Junxia Zhang,1,* Qiang Zhou,2,* Ge Gao,3,* Yanfen Wang,1 Zhihui Fang,1 Guanlin Li,4 Mengfei Yu,5 Lingfei Kong,6 Ying Xing,3 Xiaoqun Gao11Department of Anatomy, Basic Medical College, Zhengzhou University, Henan, People’s Republic of China; 2Department of Pathology, Children’s Hospital of Zhengzhou City, Henan, People’s Republic of China; 3Department of Physiology, Basic Medical College, Zhengzhou University, Henan, People’s Republic of China; 4Department of Laboratory, The First Affiliated Hospital of Zhengzhou University, Henan, People’s Republic of China; 5Department of Pharmacy, Zhengzhou Central Hospital, Zhengzhou University, Henan, People’s Republic of China; 6Department of Pathology, Henan Provincial People’s Hospital, Henan, People’s Republic of China *These authors contributed equally to this workAbstract: Glioblastoma is one of the most common malignant tumors in the nervous system in both adult and pediatric patients. Studies suggest that abnormal activation of receptor tyrosine kinases contributes to pathological development of glioblastoma. However, current therapies targeting tyrosine kinase receptors have poor therapeutic outcomes. Here, we examined anticancer effects of ponatinib, a multi-targeted tyrosine kinase inhibitor, on glioblastoma cells both in the U87MG cell line and in the mouse xenograft model. We showed that ponatinib treatment reduced cell viability and induced cell apoptosis in a dose-dependent manner in U87MG cells. In addition, ponatinib suppressed migration and invasion of U87MG cells effectively. Furthermore, ponatinib-treated tumors showed an obvious reduction of tumor volume and an increase of apoptosis as compared with vehicle-treated tumors in the mouse xenograft model. These findings support a potential application of ponatinib as a chemotherapeutic option against glioblastoma cells.Keywords: cancer therapy, glioma, glioblastoma multiforme

The whole encoding sequence for Yersinia pestis LcrV antigen was cloned into pET-32a(+) and expressed in Escherichia coli BL21 (DE3). The LcrV was high level expressed in the E. coli cytoplasm in a completely soluble form. Recombinant LcrV could be purified from the supernatant of the bacteria lysate after chromatography using a combination of Phenyl-Sepharose F F, DEAE-Sepharose F F and Hiload™ Superdex™ 75. The final yield of approximately 3 g of purified rLcrV from 42 L bioreactor containing 25 L LB medium was obtained. High-titer IgG directed against rLcrV was detected positive after immunization on the BALB/c mice. The results presented here exhibit the ability to generate multi-gram quantities of non-tagged rLcrV from E. coli that can be used for the development of vaccine for preventing plague.

Anthrax toxin consists of three proteins, protective antigen (PA), lethal factor, and edema factor. PA is the major component in the current anthrax vaccine, but the antigenic epitopes on it are not well-defined. We generated a pool of toxin-neutralizing anti-PA monoclonal antibodies (MAbs) to analyze the neutralizing epitopes of PA. Nine toxin-neutralizing MAbs obtained were found bound to three different domains of PA respectively, among which three MAbs with the strongest toxin-neutralizing activity recognized the same epitope within domain 2. This epitope was fine mapped to the chymotrypsin-sensitive site, (312)SFFD(315), in the 2beta(2)-2beta(3) loop of PA, using phage-displayed random peptide libraries and mutation analysis. The result demonstrated for the first time that the 2beta(2)-2beta(3) loop, which is involved in the transition of PA oligomers from prepore to pore, contains a dominant neutralizing epitope. This work contributes to the immunological and functional analysis of PA and offers perspective for the development of a new epitope vaccine against anthrax.