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Predicted increases in global average temperature are physiologically trivial for most endotherms. However, heat waves will also increase in both frequency and severity, and these will be physiologically more important. Lactating small mammals are hypothesized to be limited by heat dissipation capacity, suggesting high temperatures may adversely impact lactation performance. We measured reproductive performance of mice and striped hamsters (Cricetulus barabensis), including milk energy output (MEO), at temperatures between 21 and 36 °C. In both species, there was a decline in MEO between 21 and 33 °C. In mice, milk production at 33 °C was only 18% of that at 21 °C. This led to reductions in pup growth by 20% but limited pup mortality (0.8%), because of a threefold increase in growth efficiency. In contrast, in hamsters, MEO at 33 °C was reduced to 78.1% of that at 21 °C, yet this led to significant pup mortality (possibly infanticide) and reduced pup growth by 12.7%. Hamster females were more able to sustain milk production as ambient temperature increased, but they and their pups were less capable of adjusting to the lower supply. In both species, exposure to 36 °C resulted in rapid catastrophic lactation failure and maternal mortality. Upper lethal temperature was lowered by 3 to 6 °C in late lactation, making it a critically sensitive window to high ambient temperatures. Our data suggest future heat wave events will impact breeding success of small rodents, but this is based on animals with a long history in captivity. More work should be performed on wild rodents to confirm these impacts.

Three-dimensional (3D) graphene networks are performance boosters for functional nanostructures in energy-related fields. Although tremendous intriguing nanostructures-decorated 3D graphene network...

Three new species of Russulasection Ingratae, found in Guizhou and Jiangsu Provinces, southern China, are proposed: R. straminella, R. subpectinatoides and R. succinea. Photographs, line drawings and detailed morphological descriptions for these species are provided with comparisons against closely-related taxa. Phylogenetic analysis of the internal transcribed spacer (ITS) region supported the recognition of these specimens as new species. Additionally, R. indocatillus is reported for the first time from China and morphological and phylogenetic data are provided for the Chinese specimens.

National-based prospective surveillance of all-age patients with acute diarrhea was conducted in China between 2009‒2018. Here we report the etiological, epidemiological, and clinical features of the 152,792 eligible patients enrolled in this analysis. Rotavirus A and norovirus are the two leading viral pathogens detected in the patients, followed by adenovirus and astrovirus. Diarrheagenic Escherichia coli and nontyphoidal Salmonella are the two leading bacterial pathogens, followed by Shigella and Vibrio parahaemolyticus. Patients aged, Diarrhoea is a major cause of morbidity and mortality in China. Here, the authors present results from a large sentinel surveillance scheme from 217 hospitals in all 31 provinces in mainland China, including ~150,000 patients with acute diarrhoea and covering years 2009-2018.

To identify the amino acid residues that are the critical factors contributing to the high catalytic activity and good thermostability of Yersinia enterocolitica subsp. palearctica non-specific nuclease (Y. NSN), in this work, we analyzed the structure and the functional domain of Y. NSN by using bioinformatics software. Disulphide bonds were analyzed by mass spectrometry. Six active site mutants of Y. NSN, including two salt bridge mutants and four disulphide bond mutants, were obtained by site-directed mutagenesis. Their enzyme activity and thermostability were assayed. The results showed that when changing the amino acids of the active site, or breaking the salt bridges and disulphide bonds, the enzyme activity and thermal stability of Y. NSN decreased significantly, compared to those of the wild-type enzyme. It is indicated that the active site, salt bridges and disulphide bonds play an important role in the enzyme activity and thermostability of Y. NSN.

Background: To identify the critical amino acid residues that contribute to the high enzyme activity and good thermostability of Yersinia enterocolitica subsp. palearctica (Y. NSN), 15 mutants of Y. NSN were obtained by site-directed mutagenesis in this study. And their enzyme activity and thermostability were assayed. Effect of several factors on the enzyme activity and thermostability of Y. NSN, was also investigated. Results: The results showed that the I203F and D264E mutants retained approximately 75% and 70% enzyme activity, respectively, compared to the wild-type enzyme. In addition to the I203F and D264E mutants, the mutant E202A had an obvious influence on the thermostability of Y. NSN. According to the analysis of enzyme activity and thermostability of Y. NSN, we found that Glu202, Ile203 and Asp264 might be the key residues for its high enzyme activity and good thermostability. Conclusions: Among all factors affecting enzyme activity and thermostability of Y. NSN, they failed to explain the experimental results well. One reason might be that the enzyme activity and thermostability of Y. NSN were affected not only by a single factor but also by the entire environment. Normal 0 21 false false false ES X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Tabla normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin-top:0cm; mso-para-margin-right:0cm; mso-para-margin-bottom:10.0pt; mso-para-margin-left:0cm; line-height:115%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-fareast-language:EN-US;}

It has been well known that metabolic thermogenesis plays an important role in the thermoregulation of small mammals under different temperatures, while its role in fat accumulation is far from clear. In the present study, several physiological, hormonal, and biochemical measures indicative of metabolic thermogenesis were measured in the weaning striped hamsters after acclimated to a warm condition (30°C) for 1, 3 and 4months. The warm-acclimated groups significantly decreased energy intake, and simultaneously decreased nonshivering thermogenesis compared to those housed at 21°C. Body fat content increased by 29.9%, 22.1% and 19.6% in the hamsters acclimated to 1, 3 or 4months, respectively relative to their counterparts maintain at 21°C (P

// Huan Wang 1 , Fu-Ling Yan 1 , Michael Cunningham 3 , Qi-Wen Deng 1 , Lei Zuo 1 , Fang-Lan Xing 1 , Lu-Hang Shi 2 , Shan-Shan Hu 2 , Ya Huang 2 1 Neurologic Department, Affiliated Zhongda Hospital, School of Medicine, Southeast University, Nanjing, China 2 School of Medicine, Southeast University, Nanjing, China 3 Public Health School of Southeast University, Nanjing, China Correspondence to: Fu-Ling Yan, email: yanfuling218@163.com Keywords: stroke, infection, immune, sympathetic pathway, HLA-DR Received: February 25, 2016 Accepted: June 30, 2016 Published: July 08, 2016 ABSTRACT Background: Evidence has led to the consideration of immunodepression after stroke as an important contributor to stroke associated infection (SAI). However, so far no specific immunological indicator has been identified for SAI, and the underlying mechanism remains poorly understood. Results: SAI patients had significantly higher IL-6 and IL-10 levels and lower HLA-DR levels than no-infection patients within 48h after stroke onset. NA significantly increased IL-10 levels, reduced HLA-DR expression, and decreased IL-6 expression by increasing β-arrestin2 expression which reduced the activation of the NF-κB pathway. Propranolol reversed this effect of NA by reducing β-arrestin2 expression. Materials and Methods: A systematic search for eligible clinical studies was applied to pool the differences in peripheral cytokine levels between infection and no-infection stroke patients. The underlying mechanism behind these differences was investigated in vitro by applying norepinephrine (NA) and lipopolysaccharide (LPS) to simulate sympathetic pathway activation and sepsis respectively in THP-1 cells. Propranolol was applied to determine the effect of reversing the activation of the sympathetic pathway. Immunological indicators were also detected to assess the immune activation of THP-1 cells and measurements of the expression of β-arrestin2, NF-κB, IκBα and phosphor-IκBα were performed to assess the activation of the sympathetic pathway. Conclusion: IL-6, IL-10 and HLA-DR are good candidate biomarkers for SAI. The activation of the sympathetic pathway could partly account for the specific immunological alterations found in SAI patients including HLA-DR decrease and IL-10 increase, which both could be reversed by propranolol. However, the mechanism underlying IL-6 increase still needs further exploration.

Encapsulation of Enterococcus faecalis HZNU P2 ( E. faecalis HZNU P2) in the microspheres comprised of the mixtures of milk and alginate was developed in this paper. The survival of E. faecalis HZNU P2 in simulated gastric fluid (SGF, pH 2.0 and 2.5) and bile salt solution (1.0 and 2.0%) was assessed. In addition, the release and storage stability of E. faecalis HZNU P2 were studied. The results showed that encapsulated cells showed good tolerance to SGF and bile salt solution (1.0 or 2.0%). Encapsulated E. faecalis HZNU P2 could be completely released in simulated intestine fluid (SIF) within 1 h. The storage stability of encapsulated E. faecalis HZNU P2 was also highly improved. These results show that milk-alginate microspheres show a promise for the encapsulation, protection and release of E. faecalis HZNU P2 as a supplement and/or food ingredient.

Food restriction (FR) has been commonly used to decrease body fat, reducing the risk of overweight in humans and animals. However, the lost weight has been shown to be followed by overweight when food restriction ends. It remains uncertain whether the weight loss drives the overweight, or not. In the present study, striped hamsters were restricted by 15%, 30% and 40% of ad libitum food intake for 2 weeks, followed by high-fat refeeding for 6 weeks (FR15%-Re, FR30%-Re and FR40%-Re). The hamsters in FR15%, FR30% and FR40% groups decreased by 21.1%, 37.8% and 50.0% in fat mass (P 0.01), and 16.8%, 42.8% and 53.4% in leptin levels (P 0.01) compared with the hamsters fed ad libitum. The FR15%-Re, FR30%-Re and FR40%-Re groups showed 77.0%, 37.2% and 23.7% more body fat than ad libitum group (P 0.01). The FR15%-Re group showed considerable decreases in gene expression of arcuate nucleus co-expressing proopiomelanocortin (POMC), cocaine - and amphetamineregulated transcript (CART) and the long isoform of leptin receptor (LepRb) in the hypothalamus and of several genes associated with fatty acid transport to mitochondria and β-oxidation in brown adipose tissue and liver. It suggests that less weight loss is likely to drive more fat accumulation when food restriction ends, in which the impaired function of LepRb, POMC and CART in the brain and fatty acid oxidation in brown adipose tissue and liver may be involved.

Many phylogeographic studies on marine organisms in the Northwestern Pacific have supported for the biogeographic hypotheses that isolation in the marginal seas of this region during the Pleistocene glaciation lower sea level led to population genetic divergence, and thus population expansion was a common phenomenon when the sea level rebounded. However, most of these studies were based on maternally inherited mitochondrial DNA markers with limited sample sites and therefore, were unable to reveal detailed pictures encompassing paternal line information covering of the entire range. In this study, we used the mitochondrial cytochrome c oxidase subunit I (COI) and nine nuclear microsatellite loci to investigate the phylogeography of the mud snail, Bullacta exarata (Philippi, 1849), a species endemic to the Northwestern Pacific. We sampled 14 natural populations spanning across 3800 km of the Chinese coastline, essentially covering most of the species distribution range. COI analysis identified a total of 149 haplotypes separated into two distinct groups with nine mutation steps, revealing a prominent phylogeographic structure. Nuclear microsatellite data also demonstrated a similar but weaker genetic structure. The estimated time to the most recent common ancestor between the two COI haplogroups is at ∼0.89 Ma, indicating that B. exarata populations survived the Pleistocene glaciation in the Sea of Japan and the Okinawa Trough, two marginal seas around the species range. The consistent significant patterns of isolation by distance of both COI and microsatellites suggests that limited mobility of adults and short planktonic stage of larvae may have played an important role in promoting or maintaining the genetic differentiation of B. exarata. Results from population demographic analyses support population expansion late in the Pleistocene era.

To enhance the survival ability of Enterococcus faecalis HZNU P2 ( E. faecalis HZNU P2), soy protein–alginate microspheres used as the carriers were studied in this work. Characteristics of free and encapsulated cells under simulated conditions were investigated. The results so obtained showed that the viability of encapsulated cell did not decrease after 2 h incubation in simulated gastric fluid (SGF) at pH 2.5 and 2.0. Encapsulated cells experienced only minor viability reduction in bile salt solution after 1 or 2 h treatment. The viability of encapsulated cells was fully preserved after 21 days of storage at 4 °C, and only lost 0.70 and 2.0 Log CFU/g after 21 days of storage at 25 and 37 °C, respectively. Encapsulated cells could be released in simulated intestine fluid (SIF) with a fast rate. This study indicates that soy protein–alginate is a good encapsulation carrier for protecting E. faecalis HZNU P2 against adverse conditions.

The metabolic thermogenesis plays important roles in thermoregulation, and it may be also involved in body fat regulation. The thermogenesis of brown adipose tissue (BAT) is largely affected by ambient temperature, but it is unclear if the roles in body fat regulation are dependent on the temperature. In the present study, uncoupling protein 1 (ucp1)-based BAT thermogenesis, energy budget and body fat content were examined in the striped hamsters fed high fat diet (HF) at cold (5 °C) and warm (30 °C) temperatures. The effect of 2, 4-dinitrophenol (DNP), a chemical uncoupler, on body fat was also examined. The striped hamsters showed a notable increase in body fat following the HF feeding at 21 °C. The increased body fat was markedly elevated at 30 °C, but was significantly attenuated at 5 °C compared to that at 21 °C. The hamsters significantly increased energy intake at 5 °C, but consumed less food at 30 °C relative to those at 21 °C. Metabolic thermogenesis, indicated by basal metabolic rate, UCP1 expression and/or serum triiodothyronine levels, significantly increased at 5 °C, but decreased at 30 °C compared to that at 21 °C. A significant decrease in body fat content was observed in DNP-treated hamsters relative to the controls. These findings suggest that the roles of metabolic thermogenesis in body fat regulation largely depend on ambient temperature. The cold-induced enhancement of BAT thermogenesis may contribute the decreased body fat, resulting in a lean mass. Instead, the attenuation of BAT thermogenesis at the warm may result in notable obesity.

The life history of many animals includes periods of food shortage. Two behavioral strategies are involved in small mammals in response to food shortage: an increase in activity behavior, representing the increased foraging or migratory behavior, and a decrease in activity level, serving as a mechanism for conserving energy. However, it is uncertain whether animals adopt both strategies in response to food shortage, and whether hormone and neuroendocrine mechamisms are involved in both strategies. In the present study, the changes of behavior and metabolic rate were examined in food-deprived striped hamsters (Cricetulus barabensis). The effects of leptin supplement on activity behavior, metabolic rate and hypothalamic neuropeptide gene expression were also examined. The behavior of food-deprived hamsters significantly changed on basis of photoperiod phases: increasing activity during the dark phase compared to those fed ad libitum, whereas decreasing activity and simultaneously increasing resting behavior during the light phase. Resting metabolic rate, body mass, and masses of fat depots and digestive tracts significantly decreased in food-deprived hamsters compared with ad libitum controls. Leptin supplement tended to attenuate the increased activity in the dark phase. Gene expression of hypothalamic neuropeptide Y (NPY) was significantly up-regulated in food-deprived hamsters, while was significantly attenuated by exogenous leptin. These findings suggest that both behavior strategies are important behavioral adjustments in free living animals to cope with food shortage. Leptin and hypothalamic NPY gene expression may be involved in the adjustments of physiology and behavior in animals showing hyperactivity strategy in response to food shortage. This article is protected by copyright. All rights reserved

Enlarged vestibular aqueduct (EVA)‑associated hearing loss is frequently detected in individuals carrying the SLC26A4 mutation in the Chinese population. The present study aimed to identify the causative SLC26A4 coding mutations in a patient group with nonsyndromic hearing loss (NSHL) and EVA. Genomic DNA was extracted from blood samples obtained from 52 NSHL patients with EVA and from 60 normal controls. The mutation analysis for 20 coding exons of SLC26A4 was performed by direct sequencing. The results of the mutational analysis showed that there were two probands from two separate families suffering from bilateral sensorineural hearing loss with EVA, carrying the same novel compound heterozygous mutation of SLC26A4 (c.1644_1645insA and c.2168A>G). Other members of the two families had heterozygous mono‑allelic mutations with normal hearing. However, neither of these mutations were detected in the 60 normal controls. These results are the first, to the best of our knowledge, to link the compound heterozygote mutation, c.1644_1645insA and c.2168A>G, in the SLC26A4 gene to NSHL patients with EVA. The two mutations identified in the present study were located in the anti‑sigma factor antagonist domain, the core region for plasma membrane targeting of anion transporters, which suggested that the reduced or complete loss of SLC26A4 function was the direct cause of hearing loss in the two patients. These results provide a foundation for further elucidating the genetic factors responsible for EVA‑associated NSHL.

In this work, alginate-whey protein was used as wall materials for encapsulating Lactobacillus delbrueckii subsp. bulgaricus (L. bulgaricus). The characteristics of encapsulated and free L. bulgaricus showed that the free L. bulgaricus lost viability after 1 min exposure to simulated gastric fluid (SGF) at pH 2.0 and 2.5. However, the viability of encapsulated L. bulgaricus did not decrease in SGF at pH 2.5 for 2 h incubation. The viable numbers of encapsulated L. bulgaricus decreased less than 1.0 log unit for 2 h incubation in SGF at pH 2.0. For bile stability, only 1.2 log units and 2.0 log units viability of the encapsulated L. bulgaricus was lost in 1 and 2% bile for 1 h exposure, respectively, compared with no survival of free L. bulgaricus under the same conditions. Encapsulated L. bulgaricus was completely released from the microspheres in simulated intestinal fluid (SIF, pH 6.8) in 3 h. The viability of the encapsulated L. bulgaricus retained more 8.0 log CFU/g after stored at 4°C for four weeks. However, for free L. bulgaricus, only around 3.0 log CFU/mL was found at the same storage conditions. Results showed that the encapsulation could improve the stability of L. bulgaricus.

A new non-specific nuclease from Yersinia enterocolitica subsp. palearctica (Y. NSN) was expressed in Escherichia coli (E. coli) BL 21 StarTM (DE3)plysS. Induction conditions, including isopropyl-β-D-thiogalactoside (IPTG) concentration, cell density (OD600), induction time and induction temperature, were optimized using response surface methodology. Statistical analysis of the results revealed that induction temperature and all the quadratic terms of variables had significant effects on enzyme activity of Y. NSN. The optimal induction conditions were as follows: 1.5 mmol/L IPTG, OD600 of 0.80, induction time of 20.5 h, and induction temperature of 32 °C. Under the optimized conditions, the highest enzyme activity could be obtained.

Objective To develop alternative therapeutic strategy that reduces hypercholesterolemia, inflammation and atherosclerosis, we investigate if fumigaclavine C (FC), an indole alkaloid in structure, has anti-atherosclerosis function, and if so, what is the mechanism involved. Methods and results We used ApoE-deficient (ApoE −/− ) mice as an atherosclerosis model to examine if FC reduced aorta lesion size and improved serum lipid profiles. ApoE −/− mice at 6 weeks of age were fed on a western diet for 10 weeks before FC was administrated (5, 10 and 20 mg/kg) by gavage daily for additional 4 weeks. The mice were sacrificed at 20 weeks of age for examination. The atherosclerotic lesions were assessed with Oil Red O staining in the whole aorta and aortic sinus. Serum levels of triglycerides (TG), total cholesterol (TC), high density lipoprotein cholesterol (HDL-c) and low density lipoprotein cholesterol (LDL-c) were determined enzymatically. Mouse macrophages were examined for lipid droplets inside cells. FC's effect on PPARγ and PPARγ signaling pathway were further investigated by western blot and luciferase assay. We found that FC decreased atherosclerotic lesion formation in ApoE −/− mice in a dose-dependent manner. Also FC improved lipid profiles in ApoE −/− mice and reduced the foam cell numbers of peritoneal macrophages. FC stimulated PPARγ signaling pathway proteins both in vitro and in vivo. FC enhanced PPARγ transactivation activity assayed by a PPRE reporter system. Conclusion Our data indicated that FC activated PPARγ signaling pathway as well as its downstream proteins and had an effective role of anti-atherosclerosis.

In this paper, a new encapsulation carrier was applied in order to improve the survival of Lactobacillus bulgaricus ( L. bulgaricus ). L. bulgaricus was encapsulated in alginate–milk microspheres prepared by extrusion method. Around 100% encapsulation yield was achieved. The tolerance of encapsulated L. bulgaricus to adverse environments such as low pH (pH 2.0 and 2.5), high concentration of bile salt (1.0% and 2.0%) and long time storage (1 month), was investigated. Release characteristic of encapsulated L. bulgaricus in Simulated Intestine Fluid (SIF) was also studied. The results showed that encapsulation could improve the tolerance of L. bulgaricus to adverse environments. The viability of encapsulated L. bulgaricus did not change after 120 min incubation in Simulated Gastric Fluid (SGF) pH 2.5. The viability of encapsulated L. bulgaricus could be kept more than 8 log CFU/g after 120 min incubation in SGF pH 2.0. The viability of encapsulated L. bulgaricus in 1% bile salt solution was reduced from 9.98 log CFU/g microspheres to 9.24 and 8.48 log CFU/g microspheres after 1 and 2 h incubation, respectively. Around 1.3 and 2.1 log CFU/g microspheres were reduced after 1 and 2 h exposure in 2% bile salt solution, respectively. Full viability of encapsulated L. bulgaricus could be preserved after 1 month’s storage at 4 °C. L. bulgaricus encapsulated in alginate–milk microspheres could be completely released in 1 h. These studies demonstrated encapsulation of L. bulgaricus in alginate–milk microspheres is an effective protection technique against extreme simulated gastrointestinal environment.

Date fruit has served as a staple food in the Arab world for centuries. Worldwide production of date fruit has increased almost threefold over the last 40 years, reaching 7.68 million tons in 2010. Date fruit can provide many essential nutrients and potential health benefits to the consumer. Date fruit goes through four ripening stages named kimri, khalal, rutab and tamer. The main chemical components of date fruit include carbohydrates, dietary fibre, enzymes, protein, fat, minerals, vitamins, phenolic acids and carotenoids. The chemical composition of date fruit varies according to ripening stage, cultivar, growing environment, postharvest conditions, etc. The nutritional and medicinal activities of date fruit are related to its chemical composition. Many studies have shown that date fruit has antioxidant, antimutagenic, anti-inflammatory, gastroprotective, hepatoprotective, nephroprotective, anticancer and immunostimulant activities. Various date fruit-based products such as date syrup, date paste, date juice and their derived products are available. Date by-products can be used as raw materials for the production of value-added products such as organic acids, exopolysaccharides, antibiotics, date-flavoured probiotic-fermented dairy produce, bakery yeasts, etc. In this paper the chemical composition and nutritional and medicinal values of date fruit as well as date fruit-based products are reviewed.

Summary Owing to their considerable beneficial effects on human health, probiotics have been increasingly incorporated into food products. However, many findings have demonstrated that their survival and stability are very sensitive to processing and host gastrointestinal tract. To solve these problems, encapsulation techniques have been received considerable attention these days. So, in this review paper, methods for probiotics encapsulation, alginate-based and protein-based materials for probiotics encapsulation and application of encapsulated probiotics in food industry were discussed.

Lactobacillus delbrueckii subsp. bulgaricus (L. bulgaricus) was encapsulated in alginate-skim milk microspheres. Characteristics of encapsulated L. bulgaricus, such as pH stability, bile stability, storage stability and release property, were studied in this paper. The viability of free L. bulgaricus was not observed after 1 min in simulated gastric fluids (SGF) at pH 2.5 or 2.0. Compared with that of free L. bulgaricus, the viability of encapsulated L. bulgaricus only decreased 0.7 log CFU/g and 2 log CFU/g after 2.0 h incubation in SGF at pH 2.5 and pH 2.0, respectively. L. bulgaricus was also sensitive to bile solution. The viability of free L. bulgaricus was fully lost after 1 h incubation in 1 and 2% bile solution, while the viability of encapsulated L. bulgaricus was only lost 2 log CFU/g and 2.6 log CFU/g in 1 and 2% bile solution at the same time, respectively. Encapsulated L. bulgaricus could be completely released from microspheres in simulated intestinal fluid (pH 6.8) within 2 h. The viability of encapsulated L. bulgaricus retained around 8 log CFU/g when stored at 4°C for 30 days. The current encapsulation technique enables a large proportion of L. bulgaricus to remain good bioactive in a simulated gastrointestinal tract environment.

(1) To explore association between high sensitive C-reactive protein (hs-CRP) and positive rate of carotid plaques in the middle- and aged- population; (2) To analyze relationship of hs-CRP and characteristics of carotid plaques in B-mode ultrasonography. 1626 subjects (people’s republic of

Effects of various factors, such as pH, ionic strength, glutaraldehyde concentration, enzyme amount and immobilization time, on enzyme activity were investigated. The immobilization conditions were optimized by orthogonal experiments. Characterizations of immobilized nuclease p1 were also evaluated. Through orthogonal optimization, the optimal immobilization conditions were as follows: pH 5.6, ionic strength 0.125, glutaraldehyde concentration 0.20% and immobilization time 2.0 h. Optimal pH of immobilized enzyme was 5.8. Optimal temperature of immobilized enzyme was 70ºC. Thermal, operational and storage stabilities of the enzyme were improved after it was immobilized on DEAE cellulose. Michaelis constant Km of immobilized enzyme at 69ºC was found to be 27.21 g/l by the Lineweaver-Burk plot.

In this study, immobilization of nuclease p1 on paper cellulose was studied. Effect of various factors, such as etherification pH, couple pH and time, (NH4)2SO4 concentration, enzyme amount et al, on the activity of immobilized enzyme was investigated. The immobilization conditions were optimized by orthogonal experiment. Characterizations of immobilized nuclease p1 were also evaluated. Through orthogonal experiment, the optimal immobilization conditions were as follows: couple pH 12, etherification pH 7.0, (NHJ4)2SO4 concentration 10% and enzyme amount 6.0 ml. Optimal pH of immobilized enzyme was 5.8. Optimal temperature of immobilized enzyme was 80°C. Thermal, operational and storage stabilities of the enzyme were improved after it was immobilized on paper cellulose. Michaelis constant Km of immobilized enzyme was disclosed as 5.72 g/l by the Lineweaver-Burk plot at 70°C.

In this paper, production of 5′-nucleotides using a free enzyme membrane reactor was presented. The enzyme membrane reactor consists in a coupling of a membrane separation process with an enzymatic reaction. Retention of substrate (RNA) and catalyst (nuclease P1) by the ultrafiltration membranes was observed. The best enzyme membrane reactor configuration was achieved with a polyethersulfone membrane of 30 kDa molecular weight cut-off, since the substrate and biocatalyst were retained with no loss of enzyme activity. Enzyme stability and optimum temperature were investigated. Other optimized operating conditions were also obtained: working temperature 65 °C, flow rate 2.0 mL/min (transmembrane pressure 0.050 MPa), and reactor volume 100 mL. The results demonstrated that enzymatic production of 5′-nucleotides from RNA was efficiently conducted by employing an ultrafiltration membrane reactor.

The sorption of nuclease P1 onto chitosan nano-particles is studied in this paper. The effect of some adsorption kinetics factors such as nuclease P1 concentration, chitosan nano-particles solution concentration, adsorption temperature, chitosan nano-particles size, solution pH, etc. is investigated. Adsorption of nuclease P1 onto chitosan nano-particles is fitted into Lagergren first-order equation at initial nuclease P1 concentration of 3.0 mg/mL. The first-order constant for nuclease P1 is 22.98 h-1. When nuclease P1 concentration is controlled into certain region, the adsorption fits into Freundlich isothermal linear equation. A mechanism of adsorption for nuclease P1 is proposed by analyzing IR spectra. The IR spectra shows that the hydrogen bond might be the main force between the hydroxyl group, the NH2 group and the nuclease P1.

Ca2+ is a pivotal signal in human pulmonary artery smooth muscle cells (PASMCs) proliferation. Capacitative Ca2+ entry (CCE) via the store-operated channel (SOC), which encoded by the transient receptor potential (TRP) gene, is an important mechanism for regulating intracellular Ca2+ concentration ([Ca2+]i) in PASMCs. Sildenafil, a potent type 5 nucleotide-dependent phosphodiesterase (PDE) inhibitor, has been proposed as a therapeutic tool to treat or prevent pulmonary arterial hypertension (PAH); however, the mechanism of its antiproliferative effect on PASMCs remains unclear. This study was designed to investigate the possible antiproliferative mechanism of sildenafil on human PASMCs, namely, its effect on the Ca2+-signal pathway. Cultured normal PASMCs were treated with endothelin-1 (ET-1) or ET-1 plus sildenafil separately. Cell number and viability were determined with a hemocytometer or MTT assay. [Ca2+]i was measured by loading PASMCs with fura 2-AM. Expression of the TRPC1 gene and protein was detected by RT-PCR and Western blot, respectively. The results show that sildenafil dose-dependently inhibited the proliferation of PASMCs, the enhancement of basal [Ca2+]i level, increase of CCE, and upregulation of TRPC expression induced by ET-1. These results suggest that sildenafil potently inhibits ET-1–induced PASMCs proliferation and downregulation of CCE and TRPC expression may be responsible for its antiproliferative effect. Keywords:: store-operated channel, capacitative Ca2+ entry, transient receptor potential channel, pulmonary artery smooth muscle cell

The size of chitosan nano-particles had been prepared by ionic cross-linking of chitosan and tri-polyphosphate (CS-TPP), which were well dispersed and stable in aqueous solution. The physicochemical properties of nano-particles were characterized by IR spectra, SEM. Its sorption kinetics and sorption mechanism for neutral lipase were studied. The effect factors of adsorption kinetics were investigated, such as neutral lipase concentration, chitosan nano-particles solution concentration, adsorption temperature, size of chitosan nano-particles, stirring rate, solution pH, etc. Adsorption of neutral lipase on chitosan nano-particles was fitted into Lagergren first-order equation at initial neutral lipase of 3.0 mg/ml (pH 7.0). The first-order constant for neutral lipase was 23.34 h −1 . When neutral lipase concentration was controlled under certain region, it was fitted into Freundlich isothermal linear equation. Mechanism of adsorption for neutral lipase was presumed by analyzing IR spectra. The hydrogen in the carboxyl group was connected with electronegative oxygen. Electron pair was attractive to oxygen, so hydrogen atom became cation and proton. When electronegative neutral lipase was closed to chitosan nano-particle, the hydrogen bond might be formed. It was the main force between hydroxyl group, NH 2 group and neutral lipase.

Characterizations of chitosan nano-particles prepared by the ionization gelation method were discussed in this paper. The optimal pH of an immobilized enzyme was 7.5. The optimal temperature of the immobilized enzyme was 45 °C. The thermal, operational and storage stabilities of an enzyme were improved after they were immobilized on chitosan nano-particles. The chitosan nano-particles were applied to protect the activity of free neutral lipase. They could improve the enzyme activity by 13.17% than that of the free neutral lipase. The Michaelis constant Km of the immobilized enzyme was disclosed as 0.37 × 102 g/l by the Line-weaver Burk plot at 40 °C.

This work investigated the preparation of chitosan nanoparticles used as carriers for immobilized enzyme. The morphologic characterization of chitosan nanoparticles was evaluated by scanning electron microscope. The various preparation methods of chitosan nanoparticles were discussed and chosen. The effect of factors such as molecular weight of chitosan, chitosan concentration, TPP concentration, and solution pH on the size of chitosan nanoparticles was studied. Based on these results, response surface methodology was employed. The results showed that solution pH, TPP concentration, and chitosan concentration significantly affected the size of chitosan nanoparticles. The adequacy of the predictive model equation for predicting the magnitude orders of the size of chitosan nanoparticles was verified effectively by the validation data. Immobilization conditions were investigated as well. The minimum particles size was about 42 +/- 5 nm under the optimized conditions. The optimal conditions of immobilization were as follows: one milligram of neutral proteinase was immobilized on chitosan nanoparticles for about 15 min at 40 degrees C. Under the optimized conditions, the enzyme activity yield was 84.3%.

Background The objective of this paper was to study antimicrobial activity and safety of Enterococcus faecium KQ 2.6 (E. faecium KQ 2.6) isolated from peacock feces. Methods Agar well diffusion method was adopted in antimicrobial activity assay. Disk diffusion test was used to determine the antibiotic resistance. The identification and virulence potential of E. faecium KQ 2.6 were investigated using PCR amplification. Results The results indicated that cell free supernatant (CFS) of the strain had the good antimicrobial activity against selected gram-positive and gram-negative bacteria. The biochemical characteristics of antimicrobial substances were investigated. The results indicated that the antimicrobial substances were still active after treatment with catalase and proteinase, respectively. Moreover, the stability of antimicrobial substances did not change after heat treatment at 40, 50, 60, 70 and 80°C for 30 min, respectively. The activity of antimicrobial substances remained stable at 4 and −20°C after long time storage. The antimicrobial activity of CFS was compared with that of the buffer with similar strength and pH. The inhibitory zone of the buffer was apparently smaller than that of CFS, which meant that the acid in CFS was not the only factor that was contributed to antibacterial activity of CFS. The antibiotic resistance and virulence potential were evaluated using disk diffusion test and PCR amplification. The results showed that E. faecium KQ 2.6 did not harbor any tested virulence genes such as gelE, esp, asa1, cylA, efaA and hyl. It was susceptible to most of tested antibiotics except for vancomycin and polymyxin B. Conclusion E. faecium KQ 2.6 may be used as bio-preservative cultures for the production of fermented foods.

Nuclease p1 produced by Penicillium citrinum was purified and characterized. Fermentation was carried out for 49 h for optimum production. The enzyme was purified to homogeneity according to SDS-PAGE by thermal deactivation, ultrafiltration, (NH 4 ) 2 SO 4 precipitation, phenyl sepharose chromatography, ion-exchange chromatography and gel filtration. This protocol improved 93-fold purification and 5.4% recovery of enzyme activity. The molecular mass of nuclease p1 from Penicillium citrinum was estimated to be in the range from 43 to 44 kDa. The enzyme showed an obligate requirement for Zn 2+ for activity. K + had a significant effect on enzyme activity. Mg 2+ and Ca 2+ had a slight positive effect on enzyme activity. While Cu 2+ , Co 2+ , Al 3+ , SDS and Triton X-100 were strong inhibitors of nuclease p1. The enzyme was completely stable in the pH range from 5 to 6.5 and had a pH optimum of 5.4. The optimum temperature of nuclease p1 was 69 °C and enzyme was stable up to 70 °C. K m and V max for the purified enzyme were 24.28 mg/ml and 0.36 mg/ml min, respectively, with RNA as substrate.

In order to investigate the characterizations of chitosan nano-particles prepared by ionization gelation method, neutral proteinase was selected as a model enzyme to assess its potential applicability for the enzyme immobilization. The characterizations of immobilized enzyme were studied. The highest enzyme activity of immobilized enzyme was allowed at pH 7.5, identical to that of free enzyme, but the range of the optimum pH was broadened. The optimal temperature of immobilized enzyme was 55 °C, and the range of the optimum temperature was also broadened. The thermal, operational and storage stabilities of immobilized enzyme were improved after they were immobilized on chitosan nano-particles. Chitosan nano-particles were applied to protect the activity of neutral proteinase. They could improve 13.17% of neutral proteinase activity than that of free neutral proteinase at the same conditions. The Michaelis constant K m for immobilized enzyme was disclosed to 0.6629 mg/ml by Line weaver-Burk plot at 40 °C.

Cyclooxygenase-2 (cox-2) overexpression has been observed in several types of human cancers and has been implicated in carcinogenesis. To elucidate the role of cox-2 in esophageal carcinogenesis, we evaluated the expression of cox-2 in normal squamous epithelium squamous epithelial dysplasia (n=47), and squamous cell carcinoma of the esophagus (n=86) by immunohistochemistry, reverse transcription-PCR assay, and western blotting. A significant overexpression of cox-2 was observed in esophageal squamous dysplasia and squamous cell carcinoma compared with normal squamous epithelium. The immunoreactive score of cox-2 expression, an index determined by intensity and positivity of cox-2 staining, was 0.71 +/- 0.46 (mean +/- SD) in normal squamous esophagus, 2.19 +/- 1.79 in squamous epithelial dysplasia, and 2.67 +/- 1.77 in squamous cell carcinoma. The results of immunohistochemistry were confirmed by a reverse transcription-PCR assay and western blotting analysis. Cox-2 expression level was correlated with proliferation activity assessed by proliferating cell nuclear antigen (PCNA) index and MIB-1 index in dysplastic lesion (r=0.55, P0.01 with PCNA and r=0.72, P0.01 with MIB-1) and carcinoma (r=0.56, P0.01 with PCNA and r=0.72, P0.01 with MIB-1). Elevated cox-2 expression was associated with high p53 expression (p0.001) but not with clinicopathological features including age, sex, tumor size, histological grade, lymph node metastasis, and TNM stage. The results indicated that cox-2 may be involved in an early stage of squamous carcinogenesis of the esophagus, and that cox-2 overexpression was related to cell proliferation in esophageal squamous dysplasia and squamous cell carcinoma.

In this study, the method of unit sampling was adopted for two years to investigate and analyze the age structure of clonal tillers and seedlings of Agropyron michnoi and Leymus chinensis in the secondary succession process of Hulunbeier sandy vegetation in north China. The results indicate that in single and mixed communities, tillers of A. michnoi were composed of 3 or 4 age classes respectively, and those of L. chinensis were composed of 2 age classes. Seedlings of A. michnoi were all composed of 4 age classes, while those of L. chinensis were composed of 2 or 3 age classes, respectively. The tillers number of A. michnoi and L. chinensis in single community reduced by 10.8 and 29.7%, respectively in 2010 when compared with those in 2009; while in mixed community, the tillers number of A. michnoi reduced by 8.9%, but that of L. chinensis increased by 15.9%. In single community, the seedlings numbers of A. michnoi and L. chinensis, respectively reduced by 26.3 and 31.2%, while in mixed community, the seedlings number of A. michnoi reduced to 22.0%, but that of L. chinensis increased by 7.1%. The number ratio of various age classes of A. michnoi and L. chinensis seedlings in 2009 was basically the same as that of tillers in 2010. Both in single and mixed communities, the A. michnoi population tended to change from growing type to stable type or declining type, while L. chinensis population always remained increasing with slight reduction. The general tendency was obviously shown specially in mixed community. L. chinensis could be considered as the dominant species, while A. michnoi lost its dominant status and made the companion species in the community.Key words: Tillers, seedlings, clone, age structures, single community, mixed community.

In this paper, the physical and chemical characteristics, biological structure and function of a non-specific nuclease from Yersinia enterocolitica subsp. palearctica (Y. NSN) found in our group were studied using multiple bioinformatics approaches. The results showed that Y. NSN had 283 amino acids, a weight of 30,692.5ku and a certain hydrophilic property. Y. NSN had a signal peptide, no transmembrane domains and disulphide bonds. Cleavage site in Y. NSN was between pos. 23 and 24. The prediction result of the secondary structure showed Y. NSN was a coil structure-based protein. The ratio of @a-helix, @b-folded and random coil were 18.73%, 16.96% and 64.31%, respectively. Active sites were pos. 124, 125, 127, 157, 165 and 169. Mg^2^+ binding site was pos. 157. Substrate binding sites were pos. 124, 125 and 169. The analysis of multisequencing alignment and phylogenetic tree indicated that Y. NSN shared high similarity with the nuclease from Y. enterocolitica subsp. enterocolitica 8081. The enzyme activity results showed that Y. NSN was a nuclease with good thermostability.

Authorities frequently need to analyze suspicious powders and other samples for biothreat agents in order to assess environmental safety. Numerous nucleic acid detection technologies have been developed to detect and identify biowarfare agents in a timely fashion. The extraction of microbial nucleic acids from a wide variety of powdery and environmental samples to obtain a quality level adequate for these technologies still remains a technical challenge. We aimed to develop a rapid and versatile method of separating bacteria from these samples and then extracting their microbial DNA. Bacillus atrophaeus subsp. globigii was used as a simulant of Bacillus anthracis . We studied the effects of a broad variety of powdery and environmental samples on PCR detection and the steps required to alleviate their interference. With a benchmark DNA extraction procedure, 17 of the 23 samples investigated interfered with bacterial lysis and/or PCR-based detection. Therefore, we developed the d ual-filter method for a pplied r ecovery of microbial particles from e nvironmental and powdery samples (DARE). The DARE procedure allows the separation of bacteria from contaminating matrices that interfere with PCR detection. This procedure required only 2 min, while the DNA extraction process lasted 7 min, for a total of

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Numerous studies have shown that MDR1 polymorphisms in the form of single nucleotide poymorphisms or haplptype affect ciclosporin pharmacokinetics or blood concentrations in organ transplantation patients, but some results conflict with others. WHAT THIS STUDY ADDS • We had thought that the diease condition might conceal the minor effect of MDR1 polymorphisms. • We chose myasthenia gravis patients as a population in which disease conditions were less severe. • We also used different pharmacokinetics indices, such as dose-adjusted trough blood concentrations, dose-adjusted peak blood concentrations and trough blood concentrations under the same ciclosporin regimen. AIMS Ciclosporin (CsA), which is widely used in autoimmune disease and transplantation, has a narrow therapeutic index. It also shows considerable interindividual variability in its pharmacokinetics, which may be attributable to polymorphisms of the multidrug efflux pump P-glycoprotein, encoded by MDR-1. The aim was to determine the role of genetic polymorphisms in MDR-1 with respect to interindividual variability of CsA blood concentrations in myasthenia gravis (MG) patients. METHODS MG patients (n = 129) receiving CsA were genotyped for MDR-1 1236CT (exon 12), 2677GT (exon 21) and 3435CT (exon 26). Trough blood and peak blood concentrations were determined to see if there was correlation with the corresponding genotype. RESULTS We observed a trend for CsA blood concentrations, especially peak blood concentrations, to be higher with the wild-type allele compared with minor alleles in genotype and haplotype. Furthermore, under the same CsA regimen, it was found that the trough concentrations of variant genotype (ABCB1 1236TT or ABCB1 2677TT) were significant greater than those of wild-type (ABCB1 1236CC or ABCB1 2677GG, respectively) (P = 0.0222 and 0.0081). The trough concentrations of wild-type haplotype pair group were significantly lower those that of the mutant type pair group (TT-TT-TT) (P = 0.007). CONCLUSIONS ABCB1 polymorphisms in both genotype and haplotype may have a minor effect on the CsA blood concentrations.

Transient receptor potential vanilloid (TRPV) channels are nonselective cation channels pertinent to diverse physiological functions. Multiple TRPV channel subtypes have been identified in different tissues and cloned. The aim of this study was to investigate the role of TRPV channels in hypoxia-induced proliferation of human pulmonary artery smooth muscle cells (PASMCs) and its possible signal pathway. Reverse transcriptase–polymerase chain reaction, real-time polymerase chain reaction, and Western blot analysis were used to detect the expression of TRPV in human PASMCs. Cell number was determined with a hemocytometer. Cytosolic Ca2+ concentration ([Ca2+]cyt) was measured with a dynamic digital Ca2+ imaging system. The mRNA of TRPV1-4 was detected in human PASMCs and chronic hypoxia up-regulated expression levels of the TRPV1 gene and protein. The ability to proliferate, the resting [Ca2+]cyt, and cyclopiazonic acid–induced capacitative Ca2+ entry in human PASMCs were enhanced significantly by chronic hypoxia compared with the control, and these effects were inhibited in a dose-dependent manner by capsazepine, a TRPV1 channel inhibitor. These results suggest that TRPV1 may be a critical pathway or mediator in chronic hypoxia-induced proliferation of human PASMCs.

To improve the efficiency of the use of nuclease P1, enzyme immobilization technology was applied using nuclease P1. Characterization of immobilized nuclease P1 on different supports was studied. The results showed that the optimum pH and temperature of nuclease P1 immobilized via different supports were enhanced. The immobilized enzyme was obviously stable when stored for long periods and was reusable. The best results were obtained when nuclease P1 was immobilized on chitosan nanoparticles. The nanoparticles were applied to protect the activity of nuclease P1 and improved enzyme activity by 13.17% over that of free nuclease P1 at the same conditions. The Michaelis constant Km and Vmax were determined for free and immobilized enzyme as well.

To investigate the effect of X-ray repair cross complementing 1 (XRCC1) genetic polymorphisms on esophageal cancer risk, we determined XRCC1 polymorphisms at codon 194 (Arg→Trp) and codon 399 (Arg→Gln) in 135 patients with esophageal squamous cell carcinoma (ESCC) and 152 normal controls from hospitals. Although polymorphism at codon 194 was not associated with risk for ESCC, we found that the frequency of XRCC1 399 Gln/Gln genotype in ESCC patients (14.1%) was significantly higher than that in normal controls (3.3%), and that XRCC1 399 Gln/Gln genotype was associated with an increased risk of ESCC (odds ratio (OR)=5.15, 95% confidence interval (CI): 2.42–0.93). In addition, we found that the risk for smoker increased 4.2-fold than non-smokers in the 399 Gln/Gln genotype (OR=4.20, 95% CI: 2.37–7.44). These results suggest that XRCC1 399 Gln/Gln genotype may contribute to the risk of ESCC and modify risk associated with smoking.

An essential regulatory process of synaptic transmission is the inactivation of released neurotransmitters by the transmitter-specific uptake mechanism, {gamma}-Aminobutyric acid (GABA) is an inhibitory transmitter in the vertebrate central nervous system; its activity is terminated by a high-affinity Na{sup +} and Cl{sup -} -dependent specific GABA transporter (GAT), which carries the neurotransmitter to the presynaptic neuron and/or glial elements surrounding the synaptic cleft. Deficiency of the transporter may cause epilepsy and some other nervous diseases. The human GAT gene (GABATHG), approximately 25 kb in length, has been cloned and sequenced by our colleagues (7). Here the results of the in situ hybridization mapping with the gene are presented. 10 refs., 1 fig.