SKA2 promotes proliferation and invasion of hepatocellular carcinoma cells via activating the ß-catenin signaling pathway.

Spindle and kinetochore associated complex subunit 2 (SKA2) is a part of the spindle and kinetochore associated (SKA) complex, which has been reported in various cancers, including the breast cancer, lung cancer, and glioma. However, its role remains unclear in hepatocellular carcinoma (HCC). Our study found that SKA2 mRNA levels and immunohistochemistry staining were significantly increased in HCC tissues compared with normal tissues. The role of SKA2 in cell proliferation and invasion was also determined. Overexpression of SKA2 significantly promoted cell proliferation and invasion, while knocking down of SKA2 expression inhibited the growth and invasion of HCC cells. In experiments investigating the underlying mechanism, overexpression of SKA2 may increase the expression levels of total ß-catenin, and knockdown of SKA2 may decrease the expression levels of total ß-catenin. Our studies thus suggest that SKA2 may promote proliferation and invasion of hepatocellular carcinoma cells by activating the ß-catenin signaling pathway, which can serve as a potential target in the diagnosis and/or treatment of HCC.

Validation of an electronic tool for flagging surgical site infections based on clinical practice patterns for triaging surveillance: Operational successes and barriers.

BACKGROUND: Surveillance is an effective strategy for reducing surgical site infections (SSIs); however, current identification methods are resource-intensive. Therefore, we sought to validate an electronic SSI triaging tool for detection of probable infections and identify operational barriers and challenges. METHODS: A retrospective cohort study was conducted among all Veterans Affairs Surgical Quality Improvement Program (VASQIP)-reviewed surgeries at 2 Veterans Affairs medical centers from October 1, 2011-September 30, 2014. During the postoperative period, clinical and administrative variables associated with SSI (relevant microbiology order, antibiotic order, radiology order, and administrative codes) were extracted from the electronic medical record and used to score the probability (high, intermediate, and low) that an SSI occurred. VASQIP manual chart review was used as the gold standard of comparison. RESULTS: VASQIP manual review identified 118 SSIs out of 3,700 surgeries (3.2%). There were 2,041, 1,428, and 231 surgeries that met criteria for low, intermediate, and high probability for SSI. The tool's area under the curve was 0.86 (95% confidence interval, 0.82-0.89). The sensitivity among low-probability surgeries was 92.4%, and the specificity among high-probability surgeries was 95.1%. CONCLUSIONS: The electronic SSI tool has the potential to be used for triaging VASQIP surveillance toward the high-probability surgeries and to avoid manual review of surgeries with low probability of SSI.

Effects of dietary supplementation of quercetin on performance, egg quality, cecal microflora populations, and antioxidant status in laying hens.

Plant polyphenols, especially flavonoids, are of great interest due to their wide range of biological activities. Quercetin, a ubiquitous flavonoid, is known to have antioxidant and antibacterial effects. In this study, we investigated the effect of quercetin on performance, egg quality, cecal microflora populations, and antioxidant status in laying hens. Two hundred forty 28-wk-old Hessian laying hens, with an average laying rate of lay 85% at the start, were randomly allotted to 4 treatments and fed 1 of 4 diets (negative control, 0.2, 0.4, and 0.6 g of quercetin/kg of diet) for 8 wk. Layer performance responses, egg quality parameters, cecal microflora populations and antioxidant status were measured at the end of the experiment. Results showed that feed conversion decreased as the quercetin level increased. Laying rate had a quadratic correlation with the level of quercetin (P = 0.056) and was maximized by the supplementation level of 0.2 g/kg of diet. However, no significant quercetin effect was observed on egg quality. Regression analysis showed that the population of total aerobes and coliforms decreased and the population of Bifidobacteria increased as the level of quercetin increased. Regression analysis also showed the activities of Cu-Zn-superoxide dismutase increased as the level of quercetin increased (P < 0.05). Results of the study suggest that the appropriate level of supplementation is 0.367 to 0.369 g of quercetin/kg of feed based on the improvement of laying rate (with 88.55 as maximum value) and feed conversion (with 2.0725 as minimum value). Our observations provided further evidence that dietary supplementation of quercetin improved performance by modulation of intestinal environment and liver superoxide dismutase content in laying hens. Quercetin has the potential as functional feed additive in animal production.

Effect of quercetin on performance and egg quality during the late laying period of hens.

1. At the end of the peak laying period for fowl, both performance and egg quality decrease markedly. Counteracting either or both of these could, therefore, have significant positive economic implications. Quercetin, a typical representative of the flavonol compounds, has a variety of biological functions. However, there are no reported findings on its use as a feed additive. 2. In this study, we investigated the effects of quercetin on laying rate, egg quality and blood traits associated with egg quality in laying hens (Hessain) during the late laying period and explored the possibility of using quercetin as a functional feed additive. 3. The laying hens (n = 240; 39 weeks old) were randomly assigned to 4 treatments consisting of 6 replicates (n = 10) and were fed with diets containing quercetin at 0, 0.2, 0.4 or 0.6 g/kg. 4. Results showed that laying rate was increased and feed-egg ratio was decreased significantly by 0.2 and 0.4 g/kg quercetin. Compared with controls, Haugh unit, eggshell strength, eggshell thickness and yolk protein were increased, but yolk cholesterol was decreased by quercetin. 5. In conclusion, quercetin was beneficial in improving performance and egg quality. The recommended concentration of quercetin is 0.4 g/kg of the basal diet. At this concentration, increased laying rate, improved egg quality and reduced yolk cholesterol can be expected.

Identification of a missense mutation of c.3064G>A, Gly1022Ser in exon 43 of COL1A1 gene in a girl with osteogenesis imperfecta type III.

Osteogenesis imperfecta (OI) types I-V have been inherited in an autosomal dominant pattern. OI type I is associated with mutations in COL1A1 mostly due to a null allele. OI types II-IV are associated with mutations in COL1A1 or COL1A2 and mostly are due to glycine substitutions. It has been suggested that the effect of glycine substitutions is position specific, and the substitution of glycine by serine has much less lethal effect than the substitutions by valine, aspartic acid, glutamic acid, arginine and cysteine. We report identification of c.3064G>A, GGT>AGT, Gly1022Ser (Gly(844) --> Ser844 in triple helix) in exon 43 of the COL1A1 gene in an 8-year-old girl with OI type III. Our report provides evidence that at triple helix glycine residue 844 (p.Gly1022), a glycine substitution by serine can result in OI type III but not a lethal outcome.

Regulation of nucleocytoplasmic localizatioin of the atDjC6 chaperone protein.

The sequence of the atDjC6 chaperone protein includes three potential nuclear localization signal (NLS) sequences (A-C) and three potential nuclear export signal (NES) sequences (X-Z). The subcellular localization of atDjC6 was studied by scanning laser confocal microscopy of chimera with the green-fluorescent protein (GFP) transiently expressed in tobacco BY-2 cells. The localization of the atDjC6::GFP chimera was coincident with that of the nuclear stain propidium iodide. Site-directed mutagenesis was used to verify the predicted NLS sequences. Each was individually fused to GFP and tested for protein localization. The individual NLS sequences were sufficient to direct partial nuclear localization of GFP, although the targeting information within NLS-B is apparently conformation sensitive. Site-directed mutagenesis of the NES sequences increased the amount of each chimera that was nuclearly localized, indicating a decrease in nuclear export. When any pair of NLS sequences were appended to GFP, the chimera were entirely nuclearly localized. Quantitative two-hybrid analysis was used to verify that the decoding of NLS sequence information involves interaction with the NLS-receptor protein importin-alpha. Each of the NLS sequences is flanked by a site of potential Ser phosphorylation, and recombinant atDjC6 could be phosphorylated in vitro. Mutagenesis of Ser residues to the P-Ser mimic Asp interfered with nuclear targeting, apparently by preventing recognition or binding by importin-alpha. Our results are consistent with a regulated nucleocytoplasmic localization of the atDjC6 chaperone protein.

[Inhaled budesonide for severe asthma at high altitude].

OBJECTIVE: To determine the benefit of inhaled high dose budesonide combined with terbutaline in patients with severe asthma at high altitude. METHODS: 42 patients with severe asthma at high altitude were assigned in a randomized, double-blind fashion to receive either budesonide combined with terbutaline (budesonide group, 21 cases) or terbutaline with placebo (control group, 21 cases). Both groups received terbutaline delivered by a metered-dose inhaler in a dose of 2.5 mg. Budesonide delivered by a metered-dose inhaler in 1.2 mg in budesonide group and placebo managed by a specially prepared metered-dose inhaler in control group were administered after terbutaline treatment above procedures. Repeated once after 10 min. Before and after therapy, the scores of the activity of accessory respiratory muscles, dyspnea, wheezing (clinical index) and lung function were documented. RESULTS: At 1, 2, 4, 6 h after therapy, FEV1% [(43 +/- +/- 5)%, (50 +/- 5)%, (57 +/- 5)%, (67 +/- 6)%], PEF% [(47 +/- 5)%, (55 +/- 6)%, (62 +/- 7)%, (69 +/- 7)%], clinical index (5.1 +/- 0.8, 4.3 +/- 0.6, 3.5 +/- 0.6, 2.5 +/- 0.4) in budesonide group and FEV1% [(42 +/- 5)%, (44 +/- 5)%, (45 +/- 5)%, (45 +/- 5)%], PFE% [(46 +/- 5)%, (47 +/- 5)%, (49 +/- 6)%, (49 +/- 6)%], clinical index (5.3 +/- 0.7, 5.0 +/- 0.5, 4.9 +/- 0.7, 4.8 +/- 0.7) in control group were difference markedly compared with before therapy [budesonide group was (35 +/- 5)%, (38 +/- 5)%, 8.3 +/- 1.0, and control group was (33 +/- 5)%, (38 +/- 5)%, 8.3 +/- 1.1, respectively], all (P < 0.01). CONCLUSIONS: High dose of inhaled budesonide combined with terbutaline should be an effective therapy for patients with severe asthma at high altitude. Budesonide inhaled in high dose produces therapeutic effects as soon as 2 h after therapy.

[A fluorometric method for the determination of norepinephrine in tissue].

This paper reported the determination of norepinephrine (NE) by spectrofluorometry in tissue. The linear relationship between the flecoroscence intensity and the content of NE was in the range of 10-100 ng (lambdaex/lambdaem = 410 nm/505 nm). The rate of recovery is 91.68%-96.64% and RSD (n = 10) is 6.1%. This method is simple, rapid and sensitive.

[Spectrofluorometric determination of corticosterone in plasma and tissue].

Corticosterone is one of the adrenal hormones. This paper reported the determination of corticosterone in plasma and tissue of small mammals with spectrofluorometry. The linear relationship between the fluorescence intensity and the concentration of corticosterone is over the range of 0.01-0.24 microg/mL (lambdaex = 470nm, lambdaem = 525nm). The average rate of standard recovery is 96.0% and RSD (n = 10) is 4.2%. This method is simple, rapid and sensitive.

[Hydride generation-AFS determination of trace arsenic, antimony, selenium and mercury in Chinese herbal medicine in Qinghaixizang plateau].

In this paper, a hydride generation non-dispersive atomic fluorescence spectrometry method has been developed for the determination of trace arsenic, antimony, selenium and mercury in Chinese herbal medicine in Qinghai-Xizang plateau. Under the optimum condition, the quantities of fourteen interference elements are below certain non-interferent limits. The detection limits were as follow (g/mL): As 1.3, Sb 0.39, Se 0.24 and Hg 0.44. The relative standard deviation of 9 determination was As 2.4-4.2%, Sb 2.5-5.6%, Se 3.1-5.0% and Hg 3.8-5.9%. The recovery of four elements was 94.7-105.4%. This method is simple, rapid, sensitive and accurate, and has been successfully tested with As, Sb, Se and Hg in more than 20 kinds of Chinese herbal medicine in Qinghai-Xizang plateau.

Transient expression of uidA constructs in in vitro onion (Allium cepa L.) cultures following particle bombardment and Agrobacterium-mediated DNA delivery.

Particle bombardment and Agrobacterium-mediated DNA delivery into immature embryos and microbulbs were used to investigate the expression of the uidA gene in in vitro onion cultures. Both methods were successful in delivering DNA and subsequent uidA expression was observed. Optimal transient ß-glucuronidase activity was observed in immature embryos that had been pre-cultured for three days and bombarded at a distance of 3 cm from the stopping plate, under 25 in Hg vacuum, using 900-1300 psi rupture discs. The CaMV35S-uidA gene construct gave five fold higher transient ß-glucuronidase activity than the uidA gene construct regulated by any of four other promoters initially chosen for high experession in monocotyledonous tissues.

The energetic linkage of GTP hydrolysis and the synthesis of activated sulfate.

ATP sulfurylase, from Escherichia coli K-12, catalyzes both the hydrolysis of GTP and the synthesis of activated sulfate (APS). This paper describes the energetic linkage of these reactions and the events that couple them. Steady-state and single-turnover experiments suggest that the binding of GTP inhibits APS production and that the hydrolysis of GTP is required to generate the enzyme form(s) that produces APS. It is this progression from the inhibitory, E-GTP, to the productive, E-GDP, complexes in the cycle of APS synthesis that energetically links these two reactions. This model stands in contrast to other GTPase/target systems in which the binding of GTP alone is sufficient to catalyze multiple turnovers of the target reaction. The stoichiometry of GTP hydrolysis to APS synthesis is 1:1, and equilibrium measurements show that -9.1 kcal/mol, produced by the hydrolysis of GTP, is used to thermodynamically drive production of APS and PPi. These findings establish the mechanism of energy transfer in this novel GTPase/target system, and substantially alter our understanding of the energetics of sulfate activation, an essential step in the metabolic assimilation of sulfur.

The DNA sequence of the sulfate activation locus from Escherichia coli K-12.

The DNA sequence of the sulfate activation locus from Escherichia coli K-12 has been determined. The sequence includes the structural genes encoding the enzymes ATP sulfurylase (cysD and cysN) and APS kinase (cysC) which catalyze the synthesis of activated sulfate. These are the only genes known to reside in the sulfate activation operon. Consensus elements of the operon promoter were identified, and the start codons and open reading frames of the Cys polypeptides were determined. During this work, another gene, iap, was partially sequenced and mapped. The activity of ATP sulfurylase is stimulated by an intrinsic GTPase. Comparison of the primary sequences of CysN and Ef-Tu revealed that CysN has conserved many of the residues integral to the three-dimensional structure important for guanine nucleotide binding in Ef-Tu and RAS. nodP and nodQ, from Rhizobium meliloti, are essential for nodulation in leguminous plants. The Cys and Nod proteins are remarkably similar. NodP appears to be the smaller subunit of ATP sulfurylase. NodQ encodes homologues of both CysN and CysC; thus, these enzymes may be covalently associated in R. meliloti. The consensus GTP-binding sequences of NodQ and CysN are identical suggesting that NodQ encodes a regulatory GTPase.

GTPase-mediated activation of ATP sulfurylase.

GTP stimulates the synthesis of APS (adenosine 5'-phosphosulfate) by the enzyme ATP sulfurylase (ATP:sulfate adenylyltransferase, EC 2.7.7.4) via a GTPase mechanism. The activation of the enzyme, purified from Escherichia coli, is titratable with GTP. The initial rate of APS formation is increased 116-fold at a saturating concentration of GTP. The enzyme exhibits a GTPase activity that is stimulated by ATP and further enhanced by SO4; however, SO4 alone does not significantly stimulate GTP hydrolysis. The larger subunit of ATP sulfurylase, encoded by cysN, contains a GTP-binding consensus sequence common to other known GTP-binding proteins. This is the first evidence that the sulfate activation pathway is a metabolic target for regulation by a GTPase.