bluuebunny/crossref_metadata_2025 huggingface dataset:
Single Amino Acid Substitutions in HXT2.4 from Scheffersomyces stipitis Lead to Improved Cellobiose Fermentation by Engineered Saccharomyces cerevisiae
Authors: Ha Suk-Jin , Kim Heejin , Lin Yuping , Jang Myoung-Uoon , Galazka Jonathan M. , Kim Tae-Jip , Cate Jamie H. D. , Jin Yong-Su
Published: December 2012

Saccharomyces cerevisiae
cannot utilize cellobiose, but this yeast can be engineered to ferment cellobiose by introducing both cellodextrin transporter (
cdt-1
) and intracellular β-glucosidase (
gh1-1
) genes from
Neurospora crassa
. Here, we report that an engineered
S. cerevisiae
strain expressing the putative hexose transporter gene
HXT2.4
from
Scheffersomyces stipitis
and
gh1-1
can also ferment cellobiose. This result suggests that HXT2.4p may function as a cellobiose transporter when
HXT2.4
is overexpressed in
S. cerevisiae
. However, cellobiose fermentation by the engineered strain expressing
HXT2.4
and
gh1-1
was much slower and less efficient than that by an engineered strain that initially expressed
cdt-1
and
gh1-1
. The rate of cellobiose fermentation by the
HXT2.4
-expressing strain increased drastically after serial subcultures on cellobiose. Sequencing and retransformation of the isolated plasmids from a single colony of the fast cellobiose-fermenting culture led to the identification of a mutation (A291D) in HXT2.4 that is responsible for improved cellobiose fermentation by the evolved
S. cerevisiae
strain. Substitutions for alanine (A291) of negatively charged amino acids (A291E and A291D) or positively charged amino acids (A291K and A291R) significantly improved cellobiose fermentation. The mutant HXT2.4(A291D) exhibited 1.5-fold higher

K
m

and 4-fold higher
V
max
values than those from wild-type HXT2.4, whereas the expression levels were the same. These results suggest that the kinetic properties of wild-type HXT2.4 expressed in
S. cerevisiae
are suboptimal, and mutations of A291 into bulky charged amino acids might transform HXT2.4p into an efficient transporter, enabling rapid cellobiose fermentation by engineered
S. cerevisiae
strains.
Reconstitution of a Thermostable Xylan-Degrading Enzyme Mixture from the Bacterium Caldicellulosiruptor bescii
Authors: Su Xiaoyun , Han Yejun , Dodd Dylan , Moon Young Hwan , Yoshida Shosuke , Mackie Roderick I. , Cann Isaac K. O.
Published: December 2012

Xylose, the major constituent of xylans, as well as the side chain sugars, such as arabinose, can be metabolized by engineered yeasts into ethanol. Therefore, xylan-degrading enzymes that efficiently hydrolyze xylans will add value to cellulases used in hydrolysis of plant cell wall polysaccharides for conversion to biofuels. Heterogeneous xylan is a complex substrate, and it requires multiple enzymes to release its constituent sugars. However, the components of xylan-degrading enzymes are often individually characterized, leading to a dearth of research that analyzes synergistic actions of the components of xylan-degrading enzymes. In the present report, six genes predicted to encode components of the xylan-degrading enzymes of the thermophilic bacterium
Caldicellulosiruptor bescii
were expressed in
Escherichia coli
, and the recombinant proteins were investigated as individual enzymes and also as a xylan-degrading enzyme cocktail. Most of the component enzymes of the xylan-degrading enzyme mixture had similar optimal pH (5.5 to ∼6.5) and temperature (75 to ∼90°C), and this facilitated their investigation as an enzyme cocktail for deconstruction of xylans. The core enzymes (two endoxylanases and a β-xylosidase) exhibited high turnover numbers during catalysis, with the two endoxylanases yielding estimated
k
cat
values of ∼8,000 and ∼4,500 s
−1
, respectively, on soluble wheat arabinoxylan. Addition of side chain-cleaving enzymes to the core enzymes increased depolymerization of a more complex model substrate, oat spelt xylan. The
C. bescii
xylan-degrading enzyme mixture effectively hydrolyzes xylan at 65 to 80°C and can serve as a basal mixture for deconstruction of xylans in bioenergy feedstock at high temperatures.
Molecular Fingerprinting of Cyanobacteria from River Biofilms as a Water Quality Monitoring Tool
Authors: Loza Virginia , Perona Elvira , Mateo Pilar
Published: December 2012

Benthic cyanobacterial communities from Guadarrama River (Spain) biofilms were examined using temperature gradient gel electrophoresis (TGGE), comparing the results with microscopic analyses of field-fixed samples and the genetic characterization of cultured isolates from the river. Changes in the structure and composition of cyanobacterial communities and their possible association with eutrophication in the river downstream were studied by examining complex TGGE patterns, band extraction, and subsequent sequencing of 16S rRNA gene fragments. Band profiles differed among sampling sites depending on differences in water quality. The results showed that TGGE band richness decreased in a downstream direction, and there was a clear clustering of phylotypes on the basis of their origins from different locations according to their ecological requirements. Multivariate analyses (cluster analysis and canonical correspondence analysis) corroborated these differences. Results were consistent with those obtained from microscopic observations of field-fixed samples. According to the phylogenetic analysis, morphotypes observed in natural samples were the most common phylotypes in the TGGE sequences. These phylotypes were closely related toChamaesiphon,Aphanocapsa,Pleurocapsa,Cyanobium,Pseudanabaena,Phormidium, andLeptolyngbya. Differences in the populations in response to environmental variables, principally nutrient concentrations (dissolved inorganic nitrogen and soluble reactive phosphorus), were found. Some phylotypes were associated with low nutrient concentrations and high levels of dissolved oxygen, while other phylotypes were associated with eutrophic-hypertrophic conditions. These results support the view that once a community has been characterized and its genetic fingerprint obtained, this technique could be used for the purpose of monitoring rivers.
Protective Effects of Lactobacillus plantarum CCFM8610 against Acute Cadmium Toxicity in Mice
Authors: Zhai Qixiao , Wang Gang , Zhao Jianxin , Liu Xiaoming , Tian Fengwei , Zhang Hao , Chen Wei
Published: December 2012

This study evaluated the protective effects of
Lactobacillus plantarum
CCFM8610, a selected probiotic with good cadmium binding capacity, against acute cadmium toxicity in mice. Ninety mice were divided into prevention and therapy groups. In the prevention groups, CCFM8610 was administered at 10
9
CFU once daily for 7 days, followed by a single oral dose of cadmium chloride at 1.8 mg cadmium for each mouse. In the therapy groups, the same dose of CCFM8610 was administered for 2 days after an identical single dose of cadmium exposure. Mice that received neither cadmium nor culture or that received cadmium alone served as negative and positive controls, respectively. The effects of both living and dead CCFM8610 on cadmium ion concentrations in feces, liver, and kidney were determined. Moreover, the alterations in reduced glutathione (GSH), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and histopathology in the liver and kidney were investigated. The results showed that compared to the mice that received cadmium only, CCFM8610 treatment can effectively decrease intestinal cadmium absorption, reduce tissue cadmium accumulation, alleviate renal and hepatic oxidative stress, and ameliorate hepatic histopathological changes. Living CCFM8610 administered after cadmium exposure offered the most significant protection. Our results suggested that CCFM8610 is more effective against acute cadmium toxicity than a simple antioxidant treatment due to its special physiological functions and that it can be considered a new dietary therapeutic strategy against acute cadmium toxicity.
Role of the Guanine Nucleotide Exchange Factor Rom2 in Cell Wall Integrity Maintenance of Aspergillus fumigatus
Authors: Samantaray Sweta , Neubauer Michael , Helmschrott Christoph , Wagener Johannes
Published: December 2012

Aspergillus fumigatus
is a mold and the causal agent of invasive aspergillosis, a systemic disease with high lethality. Recently, we identified and functionally characterized three stress sensors implicated in the cell wall integrity (CWI) signaling of this pathogen, namely, Wsc1, Wsc3, and MidA. Here, we functionally characterize Rom2, a guanine nucleotide exchange factor with essential function for the cell wall integrity of
A. fumigatus
. A conditional
rom2
mutant has severe growth defects under repressive conditions and incorporates all phenotypes of the three cell wall integrity sensor mutants, e.g., the echinocandin sensitivity of the Δ
wsc1
mutant and the Congo red, calcofluor white, and heat sensitivity of the Δ
midA
mutant. Rom2 interacts with Rho1 and shows a similar intracellular distribution focused at the hyphal tips. Our results place Rom2 between the cell surface stress sensors Wsc1, Wsc3, MidA, and Rho1 and their downstream effector mitogen-activated protein (MAP) kinase module Bck1-Mkk2-MpkA.
MAPK-Activated Protein Kinase 2 Contributes to Clostridium difficile-Associated Inflammation
Authors: Bobo Linda D. , El Feghaly Rana E. , Chen Yee-Shiuan , Dubberke Erik R. , Han Zhuolin , Baker Alexandra H. , Li Jinmei , Burnham Carey-Ann D. , Haslam David B.
Published: December 2012

Clostridium difficile
infection (CDI) results in toxin-induced epithelial injury and marked intestinal inflammation. Fecal markers of intestinal inflammation correlate with CDI disease severity, but regulation of the inflammatory response is poorly understood. Previous studies demonstrated that
C. difficile
toxin TcdA activates p38 kinase in tissue culture cells and mouse ilium, resulting in interleukin-8 (IL-8) release. Here, we investigated the role of phosphorylated mitogen-activated protein kinase (MAPK)-activated protein kinase (MK2 kinase, pMK2), a key mediator of p38-dependent inflammation, in CDI. Exposure of cultured intestinal epithelial cells to the
C. difficile
toxins TcdA and TcdB resulted in p38-dependent MK2 activation. Toxin-induced IL-8 and GROα release required MK2 activity. We found that p38 and MK2 are activated in response to other actin-disrupting agents, suggesting that toxin-induced cytoskeleton disruption is the trigger for kinase-dependent cytokine response. Phosphorylated MK2 was detected in the intestines of
C. difficile-
infected hamsters and mice, demonstrating for the first time that the pathway is activated in infected animals. Furthermore, we found that elevated pMK2 correlated with the presence of toxigenic
C. difficile
among 100 patient stool samples submitted for
C. difficile
testing. In conclusion, we find that MK2 kinase is activated by TcdA and TcdB and regulates the expression of proinflammatory cytokines. Activation of p38-MK2 in infected animals and humans suggests that this pathway is a key driver of intestinal inflammation in patients with CDI.
Identification and Characterization of Porphyromonas gingivalis Client Proteins That Bind to Streptococcus oralis Glyceraldehyde-3-Phosphate Dehydrogenase
Authors: Maeda Kazuhiko , Nagata Hideki , Kuboniwa Masae , Ojima Miki , Osaki Tsukasa , Minamino Naoto , Amano Atsuo
Published: December 2012

Coaggregation of
Porphyromonas gingivalis
and oral streptococci is thought to play an important role in
P. gingivalis
colonization. Previously, we reported that
P. gingivalis
major fimbriae interacted with
Streptococcus oralis
glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and that amino acid residues 166 to 183 of GAPDH exhibited strong binding activity toward
P. gingivalis
fimbriae (H. Nagata, M. Iwasaki, K. Maeda, M. Kuboniwa, E. Hashino, M. Toe, N. Minamino, H. Kuwahara, and S. Shizukuishi, Infect. Immun.
77:
5130–5138, 2009). The present study aimed to identify and characterize
P. gingivalis
components other than fimbriae that interact with
S. oralis
GAPDH. A pulldown assay was performed to detect potential interactions between
P. gingivalis
client proteins and
S. oralis
recombinant GAPDH with amino acid residues 166 to 183 deleted by site-directed mutagenesis. Seven proteins, namely,
tonB
-dependent receptor protein (RagA4), arginine-specific proteinase B, 4-hydroxybutyryl-coenzyme A dehydratase (AbfD), lysine-specific proteinase, GAPDH, NAD-dependent glutamate dehydrogenase (GDH), and malate dehydrogenase (MDH), were identified by two-dimensional gel electrophoresis followed by proteomic analysis using tandem mass spectrometry. Interactions between these client proteins and
S. oralis
GAPDH were analyzed with a biomolecular interaction analysis system.
S. oralis
GAPDH showed high affinity for five of the seven client proteins (RagA4, AbfD, GAPDH, GDH, and MDH). Interactions between
P. gingivalis
and
S. oralis
were measured by a turbidimetric method and fluorescence microscopy. RagA4, AbfD, and GDH enhanced coaggregation, whereas GAPDH and MDH inhibited coaggregation. Furthermore, the expression of
luxS
in
P. gingivalis
was upregulated by RagA4, AbfD, and GDH but was downregulated by MDH. These results indicate that the five
P. gingivalis
client proteins function as regulators in
P. gingivalis
biofilm formation with oral streptococci.
Enhanced Susceptibility to Citrobacter rodentium Infection in MicroRNA-155-Deficient Mice
Authors: Clare Simon , John Victoria , Walker Alan W. , Hill Jennifer L. , Abreu-Goodger Cei , Hale Christine , Goulding David , Lawley Trevor D. , Mastroeni Pietro , Frankel Gadi , Enright Anton J. , Vigorito Elena , Dougan Gordon
Published: December 2012

MicroRNAs (miRNAs) are small noncoding molecules that control gene expression posttranscriptionally, with microRNA-155 (miR-155) one of the first to be implicated in immune regulation. Here, we show that miR-155-deficient mice are less able to eradicate a mucosal
Citrobacter rodentium
infection than wild-type C57BL/6 mice. miR-155-deficient mice exhibited prolonged colonization associated with a higher
C. rodentium
burden in gastrointestinal tissue and spread into systemic tissues. Germinal center formation and humoral immune responses against
C. rodentium
were severely impaired in infected miR-155-deficient mice. A similarly susceptible phenotype was observed in μMT mice reconstituted with miR-155-deficient B cells, indicating that miR-155 is required intrinsically for mediating protection against this predominantly luminal bacterial pathogen.
Successful treatment of massive intractable pericardial effusion in a patient with systemic lupus erythematosus with tocilizumab
Authors: Kamata Yasuyuki , Minota Seiji
Published: December 2012

A 51-year-old Japanese woman developed systemic lupus erythematosus (SLE) in 1995. In August 2005, she had massive pericardial effusion due to lupus pericarditis, which was compromising her circulation. Methylprednisolone pulse, intravenous cyclophosphamide pulse and pericardiocentesis were all ineffective. The pericardium was cut surgically to create a passage to drain the liquid into the pleural cavity. The procedure was temporarily effective; however, massive liquid accumulated in the pleural cavity within 1 year. Oral tacrolimus and topical betamethasone injection were ineffective. Since the interleukin-6 (IL-6) level in the effusion was markedly increased (1160 pg/ml), tocilizumab was administered intravenously at a dose of 8 mg/kg every 4 weeks. The effect was astonishing and only a residual amount of pericardial effusion remained. Prednisolone was tapered successfully from 15 to 5 mg daily. Tocilizumab is a treatment of choice when we confront an intractable serositis with massive effusion in SLE, if the IL-6 level is high.
Low intrinsic exercise capacity in rats predisposes to age-dependent cardiac remodeling independent of macrovascular function
Authors: Ritchie Rebecca H. , Leo Chen Huei , Qin Chengxue , Stephenson Erin J. , Bowden Marissa A. , Buxton Keith D. , Lessard Sarah J. , Rivas Donato A. , Koch Lauren G. , Britton Steven L. , Hawley John A. , Woodman Owen L.
Published: December 2012

Rats selectively bred for low (LCR) or high (HCR) intrinsic running capacity simultaneously present with contrasting risk factors for cardiovascular and metabolic disease. However, the impact of these phenotypes on left ventricular (LV) morphology and microvascular function, and their progression with aging, remains unresolved. We tested the hypothesis that the LCR phenotype induces progressive age-dependent LV remodeling and impairments in microvascular function, glucose utilization, and β-adrenergic responsiveness, compared with HCR. Hearts and vessels isolated from female LCR ( n = 22) or HCR ( n = 26) were studied at 12 and 35 wk. Nonselected N:NIH founder rats (11 wk) were also investigated ( n = 12). LCR had impaired glucose tolerance and elevated plasma insulin (but not glucose) and body-mass at 12 wk compared with HCR, with early LV remodeling. By 35 wk, LV prohypertrophic and glucose transporter GLUT4 gene expression were up- and downregulated, respectively. No differences in LV β-adrenoceptor expression or cAMP content between phenotypes were observed. Macrovascular endothelial function was predominantly nitric oxide (NO)-mediated in both phenotypes and remained intact in LCR for both age-groups. In contrast, mesenteric arteries microvascular endothelial function, which was impaired in LCR rats regardless of age. At 35 wk, endothelial-derived hyperpolarizing factor-mediated relaxation was impaired whereas the NO contribution to relaxation is intact. Furthermore, there was reduced β2-adrenoceptor responsiveness in both aorta and mesenteric LCR arteries. In conclusion, diminished intrinsic exercise capacity impairs systemic glucose tolerance and is accompanied by progressive development of LV remodeling. Impaired microvascular perfusion is a likely contributing factor to the cardiac phenotype.
The myosin motor Myo1c is required for VEGFR2 delivery to the cell surface and for angiogenic signaling
Authors: Tiwari Ajit , Jung Jae-Joon , Inamdar Shivangi M. , Nihalani Deepak , Choudhury Amit
Published: December 2012

Vascular endothelial growth factor receptor-2 (VEGFR2) is a receptor tyrosine kinase that is expressed in endothelial cells and regulates angiogenic signal transduction under both physiological and pathological conditions. VEGFR2 turnover at the plasma membrane (PM) is regulated by its transport through endocytic and secretory transport pathways. Short-range cargo trafficking along actin filaments is commonly regulated by motor proteins of myosin superfamily. In the current study, performed in primary human endothelial cells, we demonstrate that unconventional myosin 1c (Myo1c; class I family member) regulates the localization of VEGFR2 at the PM. We further demonstrate that the recruitment of VEGFR2 to the PM and its colocalization with Myo1c and caveolin-1 occur in response to VEGF-A (VEGF) stimulation. In addition, VEGF-induced delivery of VEGFR2 to the cell surface requires Myo1c; surface VEGFR2 levels are reduced in the absence of Myo1c and, more importantly, are restored by the overexpression of wild-type but not mutant Myo1c. Subcellular density gradient fractionation revealed that partitioning of VEGFR2 into caveolin-1- and Myo1c-enriched membrane fractions is dependent on VEGF stimulation. Myo1c depletion resulted in increased VEGF-induced VEGFR2 transport to the lysosomes for degradation and was rescued by applying either brefeldin A, which blocks trafficking between the endoplasmic reticulum and the Golgi complex, or dynasore, an inhibitor of dynamin-mediated endocytosis. Myo1c depletion also reduced VEGF-induced VEGFR2 phosphorylation at Y1175 and phosphorylation-dependent activation of ERK1/2 and c-Src kinase, leading to reduced cell proliferation and cell migration. This is the first report demonstrating that Myo1c is an important mediator of VEGF-induced VEGFR2 delivery to the cell surface and plays a role in angiogenic signaling.
Pressure-overload hypertrophy of the developing heart reveals activation of divergent gene and protein pathways in the left and right ventricular myocardium
Authors: Friehs Ingeborg , Cowan Douglas B. , Choi Yeong-Hoon , Black Kendra M. , Barnett Reanne , Bhasin Manoj K. , Daly Christian , Dillon Simon J. , Libermann Towia A. , McGowan Francis X. , del Nido Pedro J. , Levitsky Sidney , McCully James D.
Published: December 2012

Right ventricular (RV) and left ventricular (LV) myocardium differ in their pathophysiological response to pressure-overload hypertrophy. In this report we use microarray and proteomic analyses to identify pathways modulated by LV-aortic banding (AOB) and RV-pulmonary artery banding (PAB) in the immature heart. Newborn New Zealand White rabbits underwent banding of the descending thoracic aorta [LV-AOB; n = 6]. RV-PAB was achieved by banding the pulmonary artery ( n = 6). Controls ( n = 6 each) were sham-manipulated. After 4 (LV-AOB) and 6 (RV-PAB) wk recovery, the hearts were removed and matched RNA and proteins samples were isolated for microarray and proteomic analysis. Microarray and proteomic data demonstrate that in LV-AOB there is increased transcript expression levels for oxidative phosphorylation, mitochondria energy pathways, actin, ILK, hypoxia, calcium, and protein kinase-A signaling and increased protein expression levels of proteins for cellular macromolecular complex assembly and oxidative phosphorylation. In RV-PAB there is also an increased transcript expression levels for cardiac oxidative phosphorylation but increased protein expression levels for structural constituents of muscle, cardiac muscle tissue development, and calcium handling. These results identify divergent transcript and protein expression profiles in LV-AOB and RV-PAB and provide new insight into the biological basis of ventricular specific hypertrophy. The identification of these pathways should allow for the development of specific therapeutic interventions for targeted treatment and amelioration of LV-AOB and RV-PAB to ameliorate morbidity and mortality.
Brain Metastasis as an Initial Manifestation of Ovarian Carcinoma: A Case Report
Authors: Tuncer Zafer Selçuk , Boyraz Gokhan , Yazıcıoğlu Aslıhan , Selcuk İlker , Yücel Senem Özge
Published: December 2012

Brain metastasis is a rare and late manifestation of ovarian carcinoma. A 30-year-old woman initially presenting with neurologic symptoms was later found to have mucinous ovarian carcinoma. The patient had a 6 cm adnexal mass with multiple millimetric brain metastatic lesions. Following a surgical staging laparotomy procedure, the patient received brain irradiation and systemic chemotherapy.
Study of Syngas Conversion to Light Olefins by Response Surface Methodology
Authors: Atashi Hossein , Shiva Mehdi , Farshchi Tabrizi Farshad , Mirzaei Ali Akbar
Published: December 2012

The effect of adding MgO to a precipitated iron‐cobalt‐manganese based Fischer‐Tropsch synthesis (FTS) catalyst was investigated via response surface methodology. The catalytic performance of the catalysts was examined in a fixed bed microreactor at a total pressure of 1–7 bar, temperature of 280–380°C, MgO content of 5–25% and using a syngas having a H2 to CO ratio equal to 2.The dependence of the activity and product distribution on MgO content, temperature, and pressure was successfully correlated via full quadratic second‐order polynomial equations. The statistical analysis and response surface demonstrations indicated that MgO significantly influences the CO conversion and chain growth probability as well as ethane, propane, propylene, butylene selectivity, and alkene/alkane ratio. A strong interaction between variables was also evidenced in some cases. The decreasing effect of pressure on alkene to alkane ratio is investigated through olefin readsorption effects and CO hydrogenation kinetics. Finally, a multiobjective optimization procedure was employed to calculate the best amount of MgO content in different reactor conditions.
Molecular Pathways: Targeting Mdm2 and Mdm4 in Cancer Therapy
Authors: Li Qin , Lozano Guillermina
Published: December 2012

The p53 tumor suppressor is activated in response to cellular stresses to induce cell-cycle arrest, cellular senescence, and apoptosis. The p53 gene is inactivated by mutations in more than 50% of human tumors. In addition, tumor cells dampen p53 activities via overexpression of p53-negative regulators, in particular 2 structurally related proteins, Mdm2 and Mdm4. And yet, Mdm2 and Mdm4 possess p53-independent activities, which also contribute to tumor formation and progression. Given that Mdm2 and Mdm4 inhibit p53 activities to promote tumor development, small molecules and peptides were developed to abrogate the inhibition of p53 by Mdm proteins. Antitumor activities of these molecules have already been confirmed in preclinical studies and early-phase clinical trials. These research endeavors and clinical advances constitute the main focus of this review. Clin Cancer Res; 19(1); 34–41. ©2012 AACR.
Blocked Autophagy Using Lysosomotropic Agents Sensitizes Resistant Prostate Tumor Cells to the Novel Akt Inhibitor AZD5363
Authors: Lamoureux Francois , Thomas Christian , Crafter Claire , Kumano Masafumi , Zhang Fan , Davies Barry R. , Gleave Martin E. , Zoubeidi Amina
Published: December 2012

Purpose: Prostate cancer development is often associated with deletion or silencing of tumor suppressor phosphatase and tensin homolog (PTEN), a negative regulator of the phosphoinositide 3 kinase (PI3K)–Akt pathway, leading to resistance to various therapies in both the preclinical and clinical setting. Therefore, the PI3K–Akt pathway plays a central role in various cellular processes promoting survival signaling that can contribute to the malignant phenotype, and, consequently, is an attractive pharmacologic target. However, as single agents, the efficacy of AKT inhibitors may be limited by resistance mechanisms that result in minimal cell death in tumor cells.

Experimental Design: We investigated the effects of the Akt inhibitor AZD5363 on cell proliferation, cell cycle, apoptosis, and Akt downstream pathway proteins. Survival mechanisms induced by AZD5363 were investigated. We then examined the impacts of inhibition of autophagy in combination with AZD5363 on cell proliferation and apoptosis. Furthermore, the anticancer activity of combination treatment of the lysosomotropic inhibitor of autophagy (chloroquine) with the Akt inhibitor AZD5363 was evaluated in PC-3 prostate cancer xenografts.

Results: Here, we show that the Akt inhibitor AZD5363 affected the Akt downstream pathway by reducing p-mTOR, p-P70S6K, and p-S6K. While AZD5363 monotherapy induced G2 growth arrest and autophagy, it failed to induce significant apoptosis in PC-3 and DU145 prostate cancer cell lines. Blocking autophagy using pharmacologic inhibitors (3-methyladenine, chloroquine, and bafilomycin A) or genetic inhibitors (siRNA targeting Atg3 and Atg7) enhanced cell death induced by Akt inhibitor AZD5363 in these tumor prostate cell lines. Importantly, the combination of AZD5363 with chloroquine significantly reduced tumor volume by 84.9% compared with the control group and by 77.5% compared with either drug alone in PC3 xenografts.

Conclusion: Taken together, these data show that the Akt inhibitor AZD5363 synergizes with the lysosomotropic inhibitor of autophagy chloroquine to induce apoptosis and delay tumor progression in prostate cancer models that are resistant to monotherapy AZD5363, providing a new therapeutic approach potentially translatable to patients. Clin Cancer Res; 19(4); 833–44. ©2012 AACR.
Opposing Effects of Androgen Deprivation and Targeted Therapy on Prostate Cancer Prevention
Authors: Jia Shidong , Gao Xueliang , Lee Sang Hyun , Maira Sauveur-Michel , Wu Xiaoqiu , Stack Edward C. , Signoretti Sabina , Loda Massimo , Zhao Jean J. , Roberts Thomas M.
Published: December 2012

Prostate cancer is an ideal target for chemoprevention. To date, chemoprevention clinical trials with 5α-reductase inhibitors have yielded encouraging yet ultimately confounding results. Using a preclinical mouse model of high-grade prostatic intraepithelial neoplasia (HG-PIN) induced by PTEN loss, we observed unprecedented deteriorating effects of androgen deprivation, in which surgical castration or MDV3100 treatment accelerated disease progression of the otherwise stable HG-PIN to invasive castration-resistant prostate cancer (CRPC). As an alternative, targeting the phosphoinositide 3-kinase (PI3K) signaling pathway via either genetic ablation of genes encoding PI3K components or pharmacologic inhibition of the PI3K pathway reversed the PTEN loss–induced HG-PIN phenotype. Finally, concurrent inhibition of the PI3K and mitogen-activated protein kinase (MAPK) pathways was effective in blocking the growth of PTEN-null CRPC. Together, these data have revealed the potential adverse effects of antiandrogen chemoprevention in certain genetic contexts (such as PTEN loss) while showing the promise of targeted therapy in the clinical management of this complex and prevalent disease.

Significance: Chemoprevention with antiandrogen therapies is attractive for prostate cancer, given its prevalence and established hormonally mediated pathogenesis. However, because PTEN loss has been found in 9% to 45% of HG-PIN in the clinic, the current findings suggest that patients with PTEN-deficient prostate tumors might be better treated with PI3K-targeted therapies. Cancer Discov; 3(1); 44–51. ©2012 AACR.

This article is highlighted in the In This Issue feature, p. 1
A Systems Approach to Designing Effective Clinical Trials Using Simulations
Authors: Fusaro Vincent A. , Patil Prasad , Chi Chih-Lin , Contant Charles F. , Tonellato Peter J.
Published: December 2012

Background—

Pharmacogenetics in warfarin clinical trials have failed to show a significant benefit in comparison with standard clinical therapy. This study demonstrates a computational framework to systematically evaluate preclinical trial design of target population, pharmacogenetic algorithms, and dosing protocols to optimize primary outcomes.

Methods and Results—

We programmatically created an end-to-end framework that systematically evaluates warfarin clinical trial designs. The framework includes options to create a patient population, multiple dosing strategies including genetic-based and nongenetic clinical-based, multiple-dose adjustment protocols, pharmacokinetic/pharmacodynamics modeling and international normalization ratio prediction, and various types of outcome measures. We validated the framework by conducting 1000 simulations of the Applying Pharmacogenetic Algorithms to Individualize Dosing of Warfarin (CoumaGen) clinical trial primary end points. The simulation predicted a mean time in therapeutic range of 70.6% and 72.2% (
P
=0.47) in the standard and pharmacogenetic arms, respectively. Then, we evaluated another dosing protocol under the same original conditions and found a significant difference in the time in therapeutic range between the pharmacogenetic and standard arm (78.8% versus 73.8%;
P
=0.0065), respectively.

Conclusions—

We demonstrate that this simulation framework is useful in the preclinical assessment phase to study and evaluate design options and provide evidence to optimize the clinical trial for patient efficacy and reduced risk.
And Say the Cat Responded? Getting Closer to the Feline Gaze
Authors: White Kara
Published: December 2012

Abstract

Within the field of multispecies ethnography, a lingering question remains regarding how we can understand the nonhuman side of the human–nonhuman encounter. Many authors have ventured into this topic on a theoretical level, but none have proposed an effective methodological approach for how to achieve their goals. After examining the pitfalls experienced when acting as a volunteer at an animal shelter, I propose that in order to get closer to the feline gaze, we must first utilize an understanding of a cat’s sensory capabilities. Recognizing that a cat’s subjectivities are necessarily mediated by their bodies, understanding how they perceive the world involves a sensory experiential methodology. Highlighting the many contributions of phenomenological frameworks along with their limitations, I argue that getting closer to the feline gaze means appreciating species differences rather than arguing for the shared qualities held across species. Because of the species barrier, an interdisciplinary approach must meld phenomenological with ethological methods to grasp the interspecies relationships created by the cat–human encounter.
Impact of Bt cotton on the immune system and histology of the midgut of the fall armyworm Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae)
Authors: Cunha Franklin M. , Wanderley-Teixeira Valéria , Torres Jorge B. , Teixeira Álvaro A.C. , Alves Thiago J.S. , Brayner Fábio A.
Published: December 2012

Despite the efficiency of transgenic plants expressing Bacillus thuringiensis (Bt) toxins as insecticides against several lepidopterans, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) is one species that presents low susceptibility to most Bt crops. This study investigated the effects of the Cry1Ac toxin expressed by Bt cotton in the midgut of S. frugiperda and its effects on the humoral and cellular immune responses. Three hypotheses were proposed and tested with contributing factors for the natural tolerance of S. frugiperda: (i) midgut regenerative cells are activated by the Cry1Ac toxin, and thus renew the epithelial cells damaged by the protein, (ii) Cry1Ac increased production of nitric oxide or phenoloxidase in the hemolymph, and (iii) there are qualitative and quantitative variations in the hemocyte levels of S. frugiperda. Caterpillars were reared using Bt cotton (Acala 90B) and non-Bt isolines (Acala 90), from the first to the fourth instar. The Bt cotton promoted elongation of the epithelial cells in the midgut of S. frugiperda caterpillars. Hence, evidence only supported the hypothesised increase of phenoloxidase (ii) and qualitative and quantitative differences in hemocyte levels (iii) in insects that were fed with Bt and non-Bt cotton. These parameters seem to explain the low susceptibility of S. frugiperda to Cry1Ac toxin and they are a viable set of responses for the evaluation of other xenobiotic factors.
Body shape evolution of Chromis and Azurina species (Percifomes, Pomacentridae) of the eastern Pacific
Authors: Aguilar-Medrano Rosalía
Published: December 2012

Chromis is a circumglobal tropical and temperate genus with over 84 species of damselfishes. Studies based in osteological and molecular data have cited the relationship between Azurina and Chromis in the eastern Pacific. The main objectives of the study are: (1) to characterize size and shape in all Chromis and Azurina species of the eastern Pacific, (2) explore the phylogenetic signal of external morphology, and (3) present a hypothesis of the diversification process of this group. According to the results, there is no significant relationship between size and shape. The variation in body shape among all species is related to the height of the trunk, position of the snout and eye, and length of the caudal peduncle. The main morphologic variation between Azurina and Chromis is the degree of elongation of the body. Both Azurina species are closely related to C. punctipinnis and C. atrilobata. The morphological pattern of Azurina integrated it into Chromis. The phylogenetic pattern found by geometric morphometric analyses presented a high similarity with previous results based on molecular data. Phylogeny recovered two main clades, slender-bodied and deep-bodied species. This pattern of morphometric variation is closely related to exploitation of two different reef environments.
On Thermally Forced Circulations over Heated Terrain
Authors: Kirshbaum Daniel J.
Published: December 2012

A combination of analytical and numerical models is used to gain insight into the dynamics of thermally forced circulations over diurnally heated terrain. Solutions are obtained for two-layer flows (representing the boundary layer and the overlying free troposphere) over an isolated mountainlike heat source. A scaling based on the linearized Boussinesq system of equations is developed to quantify the strength of thermally forced updrafts and to identify three flow regimes, each with distinct dynamics and parameter sensitivities. This scaling closely matches corresponding numerical simulations in two of these regimes: the first characterized by a weakly stable boundary layer and significant background winds and the second by a strongly stable boundary layer. In the third regime, characterized by weak winds and weak boundary layer stability, this scaling is outperformed by a fundamentally different scaling based on thermodynamic heat engines. Within this regime, the inability of wind ventilation or static stability to diminish the buoyancy over the heat source leads to intense updrafts that are controlled by nonlinear dynamics. These nonlinearities create a positive feedback loop between the thermal forcing and vorticity that rapidly strengthens the circulation and contracts its central updraft into a narrow core. As the circulation intensifies under daytime heating, the warmest surface-based air is ventilated into the upper boundary layer, where it spreads laterally to occupy a broader area and, ultimately, restrain the circulation strength. The success demonstrated herein of simple theoretical models at predicting key aspects of thermally forced circulations offers hope for improved parameterization of related processes (e.g., convection initiation and aerosol venting) in large-scale models.
How Are Spring Snow Conditions in Central Canada Related to Early Warm-Season Precipitation?
Authors: Su Hua , Dickinson Robert E. , Findell Kirsten L. , Lintner Benjamin R.
Published: December 2012

The response of the warm-season atmosphere to antecedent snow anomalies has long been an area of study. This paper explores how the spring snow depth relates to subsequent precipitation in central Canada using ground observations, reanalysis datasets, and offline land surface model estimates. After removal of low-frequency ocean influences, April snow depth is found to correlate negatively with early warm-season (May–June) precipitation across a large portion of the study area. A chain of mechanisms is hypothesized to account for this observed negative relation: 1) a snow depth anomaly leads to a soil moisture anomaly, 2) the subsequent soil moisture anomaly affects ground turbulent fluxes, and 3) the atmospheric vertical structure allows dry soil to promote local convection. A detailed analysis supports this chain of mechanisms for those portions of the domain manifesting a statistically significant negative snow–precipitation correlation. For a portion of the study area, large-scale atmospheric circulation patterns also affect the early warm-season rainfall, indicating that the snow–precipitation feedback may depend on large-scale atmospheric dynamical features. This analysis suggests that spring snow conditions can contribute to warm-season precipitation predictability on a subseasonal to seasonal scale, but that the strength of such predictability varies geographically as it depends on the interplay of hydroclimatological conditions across multiple spatial scales.
The Remixing Dilemma
Authors: Hill Benjamin Mako , Monroy-Hernández Andrés
Published: December 2012

In this article we argue that there is a trade-off between generativity and originality in online communities that support open collaboration. We build on foundational theoretical work in peer production to formulate and test a series of hypotheses suggesting that the generativity of creative works is associated with moderate complexity, prominent authors, and cumulativeness. We also formulate and test three hypotheses that these qualities are associated with decreased originality in resulting derivatives. Our analysis uses a rich data set from the Scratch Online Community—a large website where young people openly share and remix animations and video games. We discuss the implications of this trade-off for the design of peer production systems that support amateur creativity.
National Supervised Toothbrushing Program and Dental Decay in Scotland
Authors: Macpherson L.M.D. , Anopa Y. , Conway D.I. , McMahon A.D.
Published: December 2012

We aimed to assess the association between the roll-out of the national nursery toothbrushing program and a reduction in dental decay in five-year-old children in a Scotland-wide population study. The intervention was supervised toothbrushing in nurseries and distribution of fluoride toothpaste and toothbrushes for home use, measured as the percentage of nurseries participating in each health service administrative board area. The endpoint was mean d3mft in 99,071 five-year-old children, covering 7% to 25% of the relevant population (in various years), who participated in multiple cross-sectional dental epidemiology surveys in 1987 to 2009. The slope of the uptake in toothbrushing was correlated with the slope in the reduction of d3mft. The mean d3mft in Years -2 to 0 (relative to that in start-up Year 0) was 3.06, reducing to 2.07 in Years 10 to 12 (difference = -0.99; 95% CI -1.08, -0.90; p < 0.001). The uptake of toothbrushing correlated with the decline in d3mft (correlation = -0.64; -0.86, -0.16; p = 0.011). The result improved when one outlying Health Board was excluded (correlation = -0.90; -0.97, -0.70; p < 0.0001). An improvement in the dental health of five-year-olds was detected and is associated with the uptake of nursery toothbrushing.
Effect of Local vs. Systemic Bisphosphonate Delivery on Dental Implant Fixation in a Model of Osteonecrosis of the Jaw
Authors: Abtahi J. , Agholme F. , Sandberg O. , Aspenberg P.
Published: December 2012

Locally applied bisphosphonates may improve the fixation of metal implants in bone. However, systemic bisphosphonate treatment is associated with a risk of osteonecrosis of the jaw (ONJ). We hypothesized that local delivery of bisphosphonate from the implant surface improves the fixation of dental implants without complications in a setting where systemic treatment induces ONJ. Forty rats were randomly allocated to 4 groups of 10. All groups received a titanium implant inserted in an extraction socket. Group I received the implants only. Group II received dexamethasone (0.5 mg/kg). Group III received dexamethasone as above plus alendronate (200 µg/kg). Group IV received zoledronate-coated implants and dexamethasone as above. The animals were sacrificed 2 weeks after tooth extraction. All 10 animals with systemic alendronate treatment developed large ONJ-like changes, while all with local treatment were completely healed. Implant removal torque was higher for the bisphosphonate-coated implants compared with the other groups ( p < 0.03 for each comparison). Micro-computed tomography of the maxilla showed more bone loss in the systemic alendronate group compared with groups receiving local treatment ( p = 0.001). Local bisphosphonate treatment appears to improve implant fixation in a setting where systemic treatment caused ONJ.
Monstrous Fears
Authors: White Joanna , Oosterhoff Pauline
Published: December 2012

Maternity for opiate users in Vietnam is challenging due to the criminalization of drug use, stigmatization, and poor specialist knowledge among health providers. This article outlines action research conducted among Vietnamese women who had experienced pregnancy while opiate-dependent. Misguided fears that their baby would be a “monstrosity” drove some women to seek out abortion. For those who chose to continue with their pregnancy, communication with health professionals was poor and misinformation concerning advised maternal practices and newborn care was common. Often women made intuitive decisions; few attempted to go “cold turkey” and withdrew suddenly from opiates while pregnant, and most new mothers chose to breastfeed—a healthy choice—despite being advised against this. The study revealed the importance of support provided outside of state services and culminated in new information for female drug users and training materials for peer counselors. The participatory process by which these outputs were developed is outlined.
A Pay-and-Stay model for tackling intruders in hybrid wireless mesh networks
Authors: Pathan Al-Sakib Khan , Abduallah Wafaa Mustafa , Khanam Shapla , Saleem Habibullah Yusuf
Published: December 2012

Intrusion handling in wireless mesh networks (WMNs) is a relatively less addressed topic. The difficulty may lie in the fact that there are other wireless networks for which some intrusion detection or prevention schemes are proposed that could also be applied in some way in a WMN setting. As those schemes are contributing, researchers may not find it necessary to specifically focus on this field. Another critical reason may be the difficulty in developing an effective scheme for WMNs. In fact, the structural differences among various wireless ad hoc networking technologies make it imperative to devise the mechanisms in subtle but critically different ways. For WMNs, there is a proper network backbone that is called mesh backbone (which is not present in many other wireless network counterparts), which supports the fringe part or the mesh clients. Hence, it is often possible to install the intrusion handling mechanisms or agents in the stable part and allowing some flexibility in the client or fringe parts. Nonetheless, instead of thinking in this pattern, we take a different approach of tackling intrusion by allowing an intruder to stay in the network as long as it proves to be worthy of staying in the network by supporting the network’s regular activities. The idea is that by not always directly purging rogue entities, one could exploit the intruder’s resources, thereby benefiting the network. We call our approach an intrusion-tackling mechanism and term it a Pay-and-Stay model. Alongside presenting the details and analysis of our model, in this paper, we also present the basics of various forms of intrusion handling in such types of networks. Using our evaluation results, we found that the model could be very effective in handling intruders and defending the network against a broad range of security attacks.
Bovine Colostrum to Children With Short Bowel Syndrome
Authors: Aunsholt Lise , Jeppesen Palle Bekker , Lund Pernille , Sangild Per Torp , Ifaoui Inge Bøtker Rasmussen , Qvist Niels , Husby Steffen
Published: December 2012

Background: Management of short bowel syndrome (SBS) aims to achieve intestinal autonomy to prevent fluid, electrolyte, and nutrient deficiencies and maintain adequate development. Remnant intestinal adaptation is required to obtain autonomy. In the newborn pig, colostrum has been shown to support intestinal development and hence adaptive processes. Aim: The efficacy of bovine colostrum to improve intestinal function in children with SBS was evaluated by metabolic balance studies. Materials and Methods: Nine children with SBS were included in a randomized, double‐blind, crossover study. Twenty percent of enteral fluid intake was replaced with bovine colostrum or a mixed milk diet for 4 weeks, separated by a 4‐week washout period. Intestinal absorption of energy and wet weight was used to assess intestinal function and the efficacy of colostrum. Results: Colostrum did not improve energy or wet weight absorption compared with the mixed milk diet (P = 1.00 and P = .93, respectively). Growth as measured by weight and knemometry did not differ between diets (P = .93 and P = .28). In these patients, <150% enteral energy absorption of basal metabolic rate and 50% enteral fluid absorption of basal fluid requirement suggested intestinal failure and a need for parenteral nutrition (PN). Conclusion: Inclusion of bovine colostrum to the diet did not improve intestinal function. Metabolic nutrient and wet weight balance studies successfully assessed intestinal function, and this method may distinguish between intestinal insufficiency (non–PN‐dependent) and intestinal failure (PN‐dependent) patients.