bluuebunny/crossref_metadata_2025 huggingface dataset:
Tissue factor and glycoprotein C on herpes simplex virus type 1 are protease-activated receptor 2 cofactors that enhance infection
Authors: Sutherland Michael R. , Ruf Wolfram , Pryzdial Edward L. G.
Published: February 2012

The coagulation system provides physiologic host defense, but it can also be exploited by pathogens for infection. On the HSV1 surface, host-cell–derived tissue factor (TF) and virus-encoded glycoprotein C (gC) can stimulate protease activated receptor 1 (PAR1)–enhanced infection by triggering thrombin production. Using novel engineered HSV1 variants deficient in either TF and/or gC, in the present study, we show that activated coagulation factors X (FXa) or VII (FVIIa) directly affect HSV1 infection of human umbilical vein endothelial cells in a manner that is dependent on viral TF and gC. The combination of FXa and FVIIa maximally enhanced infection for TF+/gC+ HSV1 and receptor desensitization and Ab inhibition demonstrated that both proteases act on PAR2. Inhibitory TF Abs showed that the required TF source was viral. Individually, TF or gC partly enhanced the effect of FXa, but not FVIIa, revealing gC as a novel PAR2 cofactor for FVIIa. In sharp contrast, thrombin enhanced infection via PAR1 independently of viral TF and gC. Thrombin combined with FXa/FVIIa enhanced infection, suggesting that PAR1 and PAR2 are independently involved in virus propagation. These results show that HSV1 surface cofactors promote cellular PAR2-mediated infection, indicating a novel mode by which pathogens exploit the initiation phase of the host hemostatic system.
Role of minimal residual disease monitoring in acute promyelocytic leukemia treated with arsenic trioxide in frontline therapy
Authors: Chendamarai Ezhilarasi , Balasubramanian Poonkuzhali , George Biju , Viswabandya Auro , Abraham Aby , Ahmed Rayaz , Alex Ansu Abu , Ganesan Saravanan , Lakshmi Kavitha M. , Sitaram Usha , Nair Sukesh Chandran , Chandy Mammen , Janet Nancy Beryl , Srivastava Vivi M. , Srivastava Alok , Mathews Vikram
Published: February 2012

Data on minimal residual disease (MRD) monitoring in acute promyelocytic leukemia (APL) are available only in the context of conventional all-trans retinoic acid plus chemotherapy regimens. It is recognized that the kinetics of leukemia clearance is different with the use of arsenic trioxide (ATO) in the treatment of APL. We undertook a prospective peripheral blood RT-PCR–based MRD monitoring study on patients with APL treated with a single agent ATO regimen. A total of 151 patients were enrolled in this study. A positive RT-PCR reading at the end of induction therapy was significantly associated on a multivariate analysis with an increased risk of relapse (relative risk = 4.9; P = .034). None of the good risk patients who were RT-PCR negative at the end of induction relapsed. The majority of the relapses (91%) happened within 3 years of completion of treatment. After achievement of molecular remission, the current MRD monitoring strategy was able to predict relapse in 60% of cases with an overall sensitivity and specificity of 60% and 93.2%, respectively. High-risk group patients and those that remain RT-PCR positive at the end of induction are likely to benefit from serial MRD monitoring by RT-PCR for a period of 3 years from completion of therapy.
Distinct but phenotypically heterogeneous human cell populations produce rapid recovery of platelets and neutrophils after transplantation
Authors: Cheung Alice M. S. , Leung Donna , Rostamirad Shabnam , Dhillon Kiran , Miller Paul H. , Droumeva Radina , Brinkman Ryan R. , Hogge Donna , Roy Denis Claude , Eaves Connie J.
Published: February 2012

Delayed recovery of mature blood cells poses a serious, expensive, and often life-threatening problem for many stem cell transplantation recipients, particularly if heavily pretreated and serving as their own donor, or having a CB transplantation as the only therapeutic option. Importantly, the different cells required to ensure a rapid, as well as a permanent, hematopoietic recovery in these patients remain poorly defined. We now show that human CB and mobilized peripheral blood (mPB) collections contain cells that produce platelets and neutrophils within 3 weeks after being transplanted into sublethally irradiated NOD/scid-IL-2Rγc-null mice. The cells responsible for these 2 outputs are similarly distributed between the aldehyde dehydrogenase–positive and –negative subsets of lineage marker-negative CB and mPB cells, but their overall frequencies vary independently in individual samples. In addition, their total numbers can be seen to be much (> 30-fold) lower in a single “average” CB transplantation compared with a single “average” mPB transplantation (normalized for a similar weight of the recipient), consistent with the published differential performance in adult patients of these 2 transplantation products. Experimental testing confirmed the clinical relevance of the surrogate xenotransplantation assay for quantifying cells with rapid platelet regenerative activity, underscoring its potential for future applications.
Spatial memory decline after masticatory deprivation and aging is associated with altered laminar distribution of CA1 astrocytes
Authors: Frota de Almeida Marina Negrão , de Siqueira Mendes Fabíola de Carvalho Chaves , Gurgel Felício André Pinheiro , Falsoni Manoela , Ferreira de Andrade Márcia Lorena , Bento-Torres João , da Costa Vasconcelos Pedro Fernando , Perry Victor Hugh , Picanço-Diniz Cristovam Wanderley , Kronka Sosthenes Marcia Consentino
Published: February 2012

Background

Chewing imbalances are associated with neurodegeneration and are risk factors for senile dementia in humans and memory deficits in experimental animals. We investigated the impact of long-term reduced mastication on spatial memory in young, mature and aged female albino Swiss mice by stereological analysis of the laminar distribution of CA1 astrocytes. A soft diet (SD) was used to reduce mastication in the experimental group, whereas the control group was fed a hard diet (HD). Assays were performed in 3-, 6- and 18-month-old SD and HD mice.

Results

Eating a SD variably affected the number of astrocytes in the CA1 hippocampal field, and SD mice performed worse on water maze memory tests than HD mice. Three-month-old mice in both groups could remember/find a hidden platform in the water maze. However, 6-month-old SD mice, but not HD mice, exhibited significant spatial memory dysfunction. Both SD and HD 18-month-old mice showed spatial memory decline. Older SD mice had astrocyte hyperplasia in the strata pyramidale and oriens compared to 6-month-old mice. Aging induced astrocyte hypoplasia at 18 months in the lacunosum-moleculare layer of HD mice.

Conclusions

Taken together, these results suggest that the impaired spatial learning and memory induced by masticatory deprivation and aging may be associated with altered astrocyte laminar distribution and number in the CA1 hippocampal field. The underlying molecular mechanisms are unknown and merit further investigation.
Recombinant production of the therapeutic peptide lunasin
Authors: Kyle Stuart , James Kier AR , McPherson Michael J
Published: February 2012

Background

Lunasin is a chemopreventive peptide produced in a number of plant species. It comprises a helical region with homology to a region of chromatin binding proteins, an Arg-Gly-Asp cell adhesion motif and eight aspartic acid residues. In vitro studies indicate that lunasin suppresses chemical and oncogene driven transformation of mammalian cells. We have explored efficient recombinant production of lunasin by exploiting the Clostridium thermocellum CipB cellulose binding domain (CBD) as a fusion partner protein.

Results

We used a pET28 vector to express a CBD-lunasin fusion with a hexahistidine tag and Tobacco Etch Virus protease site, to allow protease-mediated release of native lunasin. Autoinduction in E. coli BL21 (DE3) Star cells achieved expression of 3.35 g/L of CBD-lunasin fusion protein. The final yield of lunasin was 210 mg/L corresponding to 32% of the theoretical yield. Purification by cellulose binding and nickel affinity chromatography were tested with the latter proving more satisfactory.

The effects of CBD-lunasin expression on growth and morphology of the E. coli cells were examined by light and electron microscopy revealing an altered morphology in a proportion of cells. Cell division appeared to be inhibited in these cells resulting in elongated, non-septated cells.

Conclusions

The use of CBD as a fusion partner gave high protein yields by autoinduction, with lunasin release by TEV protease cleavage. With some optimisation this approach could provide a potentially valuable route for production of this therapeutic peptide. Over-expression in the host cells manifest as a cell division defect in a population of the cells, presumably mimicking some aspect of the chemopreventive function observed in mammalian cells.
Comparative studies on single-layer reduced graphene oxide films obtained by electrochemical reduction and hydrazine vapor reduction
Authors: Wang Zhijuan , Wu Shixin , Zhang Juan , Chen Peng , Yang Guocheng , Zhou Xiaozhu , Zhang Qichun , Yan Qingyu , Zhang Hua
Published: February 2012

The comparison between two kinds of single-layer reduced graphene oxide (rGO) sheets, obtained by reduction of graphene oxide (GO) with the electrochemical method and hydrazine vapor reduction, referred to as E-rGO and C-rGO, respectively, is systematically studied. Although there is no morphology difference between the E-rGO and C-rGO films adsorbed on solid substrates observed by AFM, the reduction process to obtain the E-rGO and C-rGO films is quite different. In the hydrazine vapor reduction, the nitrogen element is incorporated into the obtained C-rGO film, while no additional element is introduced to the E-rGO film during the electrochemical reduction. Moreover, Raman spectra show that the electrochemical method is more effective than the hydrazine vapor reduction method to reduce the GO films. In addition, E-rGO shows better electrocatalysis towards dopamine than does C-rGO. This study is helpful for researchers to understand these two different reduction methods and choose a suitable one to reduce GO based on their experimental requirements.
A junctionless SONOS nonvolatile memory device constructed with in situ-doped polycrystalline silicon nanowires
Authors: Su Chun-Jung , Su Tuan-Kai , Tsai Tzu-I , Lin Horng-Chih , Huang Tiao-Yuan
Published: February 2012

In this paper, a silicon-oxide-nitride-silicon nonvolatile memory constructed on an n+-poly-Si nanowire [NW] structure featuring a junctionless [JL] configuration is presented. The JL structure is fulfilled by employing only one in situ heavily phosphorous-doped poly-Si layer to simultaneously serve as source/drain regions and NW channels, thus greatly simplifying the manufacturing process and alleviating the requirement of precise control of the doping profile. Owing to the higher carrier concentration in the channel, the developed JL NW device exhibits significantly enhanced programming speed and larger memory window than its counterpart with conventional undoped-NW-channel. Moreover, it also displays acceptable erase and data retention properties. Hence, the desirable memory characteristics along with the much simplified fabrication process make the JL NW memory structure a promising candidate for future system-on-panel and three-dimensional ultrahigh density memory applications.
Compartmentation of glycogen metabolism revealed from 13C isotopologue distributions
Authors: Marin de Mas Igor , Selivanov Vitaly A , Marin Silvia , Roca Josep , Orešič Matej , Agius Loranne , Cascante Marta
Published: February 2012

Background

Stable isotope tracers are used to assess metabolic flux profiles in living cells. The existing methods of measurement average out the isotopic isomer distribution in metabolites throughout the cell, whereas the knowledge of compartmental organization of analyzed pathways is crucial for the evaluation of true fluxes. That is why we accepted a challenge to create a software tool that allows deciphering the compartmentation of metabolites based on the analysis of average isotopic isomer distribution.

Results

The software Isodyn, which simulates the dynamics of isotopic isomer distribution in central metabolic pathways, was supplemented by algorithms facilitating the transition between various analyzed metabolic schemes, and by the tools for model discrimination. It simulated 13C isotope distributions in glucose, lactate, glutamate and glycogen, measured by mass spectrometry after incubation of hepatocytes in the presence of only labeled glucose or glucose and lactate together (with label either in glucose or lactate). The simulations assumed either a single intracellular hexose phosphate pool, or also channeling of hexose phosphates resulting in a different isotopic composition of glycogen. Model discrimination test was applied to check the consistency of both models with experimental data. Metabolic flux profiles, evaluated with the accepted model that assumes channeling, revealed the range of changes in metabolic fluxes in liver cells.

Conclusions

The analysis of compartmentation of metabolic networks based on the measured 13C distribution was included in Isodyn as a routine procedure. The advantage of this implementation is that, being a part of evaluation of metabolic fluxes, it does not require additional experiments to study metabolic compartmentation. The analysis of experimental data revealed that the distribution of measured 13C-labeled glucose metabolites is inconsistent with the idea of perfect mixing of hexose phosphates in cytosol. In contrast, the observed distribution indicates the presence of a separate pool of hexose phosphates that is channeled towards glycogen synthesis.
Erythropoietin prevents PC12 cells from beta-amyloid-induced apoptosis via PI3K⁄Akt pathway
Authors: Zhi-Kun Sun , Hong-Qi Yang , Zhi-Quan Wang , Jing Pan , Zhen Hong , Sheng-Di Chen
Published: February 2012

Background

Several studies indicated that Erythropoietin (Epo) may provide remarkable neuroprotection in some neurological diseases. It also showed the significant decrease of Epo immunoreactivity in the cerebral cortex and hippocampus in aged rats, suggesting the role of Epo in the pathogenesis of age-related neurodegenerative diseases such as AD.

Methods

The protective effect of Epo was studied in differentiated PC12 cells treated with Abeta. The viability of the cells, the apoptosis of the cells and the level of Bax, Bcl-2, cleaved caspase-3 and cleaved PARP expression were detected by MTT, Hoechst 33258 staining and Western blotting respectively.

Results

20 μM Abeta (25-35) could induce a decreased viability and a increased apoptosis in PC12 cell in a time-dependent manner. However, 20 μM Abeta (35-25) had no effect on cell viability and apoptosis. Western blot analysis also showed that Abeta(25-35) treatment could decrease the expression of Bcl-2 (P < 0.05) and increase the expression of Bax (P < 0.05), Cleaved casapase-3 (P < 0.05), and Cleaved PARP (P < 0.05). The pretreatment of Epo could effectively reverse all the above changes induced by Abeta(25-35) (P < 0.05). Furthermore, the protective effect of Epo could be blocked by PI3K inhibitor LY294002 (P < 0.05).

Conclusions

Epo prevented cell injuries in PC12 cells exposed to the Abeta(25-35) and this effect may depend on the PI3K⁄Akt pathway. Our study provided an important evidence for the potential application of Epo in the therapy of Alzheimer's disease.
High level expression of bikunin in Pichia pastoris by fusion of human serum albumin
Authors: Gou Xing-Hua , Liu Yu-Ying , Chen Qi-Lei , Tang Jian-Jun , Liu Da-Yu , Zou Liang , Wu Xiao-Yong , Wang Wei
Published: February 2012

Bikunin is a proteoglycan exhibiting broad-spectrum inhibitory activity against serine proteases and could potentially suppress tumor cell invasion and metastasis. Here, we have successfully expressed recombinant human bikunin (rh-bikunin) in Pichia pastoris and also established the purification procedure. Different fusion genes of h-UTI and domain I, domain I and domain II, domain I, domain II and domain III of human serum albumin (HSA) were inserted into expression vector pPICZαA. After expressed in shake flask, rh-bikunin was produced in an 30-L fermenter and purified by affinity chromatography and cation exchange chromatography. The final expression levels were 200 mg/L and we got totally 1.08 g (3650 IU/mg) of active purified rh-bikunin (purity is 98%) from 20 L of fermentation broth. The rh-bikunin consists of unique form with molecular masses of 25 kDa, and has the same N-terminals sequence as human native bikunin. This study provided a new method for high level expression of active rh-bikunin by using HSA as fusion parter.
Metabolic Disorders and Adipose Tissue Insulin Responsiveness in Neonatally STZ-Induced Diabetic Rats Are Improved by Long-Term Melatonin Treatment
Authors: de Oliveira Ariclécio C. , Andreotti Sandra , Farias Talita da S.M. , Torres-Leal Francisco L. , de Proença André R.G. , Campaña Amanda B. , de Souza Arnaldo H. , Sertié Rogério A.L. , Carpinelli Ângelo R. , Cipolla-Neto José , Lima Fábio B.
Published: February 2012

Diabetes mellitus is a product of low insulin sensibility and pancreatic β-cell insufficiency. Rats with streptozotocin-induced diabetes during the neonatal period by the fifth day of age develop the classic diabetic picture of hyperglycemia, hypoinsulinemia, polyuria, and polydipsia aggravated by insulin resistance in adulthood. In this study, we investigated whether the effect of long-term treatment with melatonin can improve insulin resistance and other metabolic disorders in these animals. At the fourth week of age, diabetic animals started an 8-wk treatment with melatonin (1 mg/kg body weight) in the drinking water at night. Animals were then killing, and the sc, epididymal (EP), and retroperitoneal (RP) fat pads were excised, weighed, and processed for adipocyte isolation for morphometric analysis as well as for measuring glucose uptake, oxidation, and incorporation of glucose into lipids. Blood samples were collected for biochemical assays. Melatonin treatment reduced hyperglycemia, polydipsia, and polyphagia as well as improved insulin resistance as demonstrated by constant glucose disappearance rate and homeostasis model of assessment-insulin resistance. However, melatonin treatment was unable to recover body weight deficiency, fat mass, and adipocyte size of diabetic animals. Adiponectin and fructosamine levels were completely recovered by melatonin, whereas neither plasma insulin level nor insulin secretion capacity was improved in diabetic animals. Furthermore, melatonin caused a marked delay in the sexual development, leaving genital structures smaller than those of nontreated diabetic animals. Melatonin treatment improved the responsiveness of adipocytes to insulin in diabetic animals measured by tests of glucose uptake (sc, EP, and RP), glucose oxidation, and incorporation of glucose into lipids (EP and RP), an effect that seems partially related to an increased expression of insulin receptor substrate 1, acetyl-coenzyme A carboxylase and fatty acid synthase. In conclusion, melatonin treatment was capable of ameliorating the metabolic abnormalities in this particular diabetes model, including insulin resistance and promoting a better long-term glycemic control.
Systemic Leptin Administration in Supraphysiological Doses Maintains Bone Mineral Density and Mechanical Strength Despite Significant Weight Loss
Authors: Stunes Astrid K. , Westbroek Irene , Gordeladze Jan O. , Gustafsson Björn I. , Reseland Janne E. , Syversen Unni
Published: February 2012

The effects of leptin on bone are controversial. Although in vitro studies have shown that leptin stimulates osteoblast differentiation and mineralization and inhibits osteoclastogenesis, some rodent studies have shown that leptin administered centrally might result in decreased bone formation. In the present study we have investigated the skeletal effects of supraphysiological concentrations of leptin administered sc to rats. Female Fischer rats were given leptin 100 μg/d, 200 μg/d, or saline by continuous infusion for 9 wk. Bone mineral density (BMD) was measured by dual energy x-ray absorptiometry, bone microarchitecture was analyzed by micro-computed tomography, and biomechanical properties were tested by three-point bending experiments. At the end of the study, the body weight was significantly lower in rats receiving leptin compared with controls (−10.8% and −12.0% in low- and high-dose leptin groups, respectively). The high-dose leptin group also significantly lost weight compared with baseline. The plasma leptin concentration was 14- and 33-fold increased in the low- and high-dose groups, respectively. No significant differences in femoral BMD were observed. Whole-body BMD was significantly lower in the low-dose leptin group, whereas there was no difference between the high-dose leptin group and the control. Mechanical strength and microarchitecture were similar in the high-dose and the control group. The low-dose group, however, had decreased cortical volume in the femoral metaphysis, lowered bone strength, and altered moment of inertia. In conclusion, leptin given at very high doses maintains BMD, microarchitecture, and mechanical strength in female rats, despite a significant decrease in body weight.
Hematopoetic Prostaglandin D Synthase: An ESR1-Dependent Oviductal Epithelial Cell Synthase
Authors: Bridges Phillip J. , Jeoung Myoungkun , Shim Sarah , Park Ji Yeon , Lee Jae Eun , Sapsford Lindsay A. , Trudgen Kourtney , Ko Chemyong , Gye Myung Chan , Jo Misung
Published: February 2012

Oviductal disease is a primary cause of infertility, a problem that largely stems from excessive inflammation of this key reproductive organ. Our poor understanding of the mechanisms regulating oviductal inflammation restricts our ability to diagnose, treat, and/or prevent oviductal disease. Using mice, our objective was to determine the spatial localization, regulatory mechanism, and functional attributes of a hypothesized regulator of oviductal inflammation, the hematopoietic form of prostaglandin D synthase (HPGDS). Immunohistochemistry revealed specific localization of HPGDS to the oviduct's epithelium. In the isthmus, expression of HPGDS was consistent. In the ampulla, expression of HPGDS appeared dependent upon stage of the estrous cycle. HPGDS was expressed in the epithelium of immature and cycling mice but not in the oviducts of estrogen receptor α knockouts. Two receptor subtypes bind PGD2: PGD2 receptor and G protein-coupled receptor 44. Expression of mRNA for Ptgdr was higher in the epithelial cells (EPI) than in the stroma (P < 0.05), whereas mRNA for Gpr44 was higher in the stroma than epithelium (P < 0.05). Treatment of human oviductal EPI with HQL-79, an inhibitor of HPGDS, decreased cell viability (P < 0.05). Treatment of mice with HQL-79 increased mRNA for chemokine (C-C motif) ligands 3, 4, and 19; chemokine (C-X-C motif) ligands 11 and 12; IL-13 and IL-17B; and TNF receptor superfamily, member 1b (P < 0.02 for each mRNA). Overall, these results suggest that HPGDS may play a role in the regulation of inflammation and EPI health within the oviduct.
Gonadotropin-Inhibitory Hormone Inhibits GnRH-Induced Gonadotropin Subunit Gene Transcriptions by Inhibiting AC/cAMP/PKA-Dependent ERK Pathway in LβT2 Cells
Authors: Son You Lee , Ubuka Takayoshi , Millar Robert P. , Kanasaki Haruhiko , Tsutsui Kazuyoshi
Published: February 2012

A neuropeptide that directly inhibits gonadotropin secretion from the pituitary was discovered in quail and named gonadotropin-inhibitory hormone (GnIH). The presence and functional roles of GnIH orthologs, RF-amide-related peptides (RFRP), that possess a common C-terminal LPXRF-amide (X = L or Q) motif have also been demonstrated in mammals. GnIH orthologs inhibit gonadotropin synthesis and release by acting on pituitary gonadotropes and GnRH neurons in the hypothalamus via its receptor (GnIH receptor). It is becoming increasingly clear that GnIH is an important hypothalamic neuropeptide controlling reproduction, but the detailed signaling pathway mediating the inhibitory effect of GnIH on target cells is still unknown. In the present study, we investigated the pathway of GnIH cell signaling and its possible interaction with GnRH signaling using a mouse gonadotrope cell line, LβT2. First, we demonstrated the expression of GnIH receptor mRNA in LβT2 cells by RT-PCR. We then examined the inhibitory effects of mouse GnIH orthologs [mouse RFRP (mRFRP)] on GnRH-induced cell signaling events. We showed that mRFRP effectively inhibited GnRH-induced cAMP signaling by using a cAMP-sensitive reporter system and measuring cAMP levels, indicating that mRFRP function as an inhibitor of adenylate cyclase. We further showed that mRFRP inhibited GnRH-stimulated ERK phosphorylation, and this effect was mediated by the inhibition of the protein kinase A pathway. Finally, we demonstrated that mRFRP inhibited GnRH-stimulated gonadotropin subunit gene transcriptions and also LH release. Taken together, the results indicate that mRFRP function as GnIH to inhibit GnRH-induced gonadotropin subunit gene transcriptions by inhibiting adenylate cyclase/cAMP/protein kinase A-dependent ERK activation in LβT2 cells.
Transcriptome Analysis Reveals Differentially Expressed Transcripts in Rat Adrenal Zona Glomerulosa and Zona Fasciculata
Authors: Nishimoto Koshiro , Rigsby Christine S. , Wang Tao , Mukai Kuniaki , Gomez-Sanchez Celso E. , Rainey William E. , Seki Tsugio
Published: February 2012

In mammals, aldosterone is produced in the zona glomerulosa (zG), the outermost layer of the adrenal cortex, whereas glucocorticoids are produced in adjacent zona fasciculata (zF). However, the cellular mechanisms controlling the zonal development and the differential hormone production (i.e. functional zonation) are poorly understood. To explore the mechanisms, we defined zone-specific transcripts in this study. Eleven-week-old male rats were used and adrenal tissues were collected from zG and zF using laser-capture microdissection. RNA was isolated, biotin labeled, amplified, and hybridized to Illumina microarray chips. The microarray data were compared by fold change calculations. In zG, 235 transcripts showed more than a 2-fold up-regulation compared to zF with statistical significance. Similarly, 231 transcripts showed up-regulation in zF. The microarray findings were validated using quantitative RT-PCR and immunohistochemical staining on selected transcripts, including Cyp11b2 (zG/zF: 214.2x), Rgs4 (68.4x), Smoc2 (49.3x), and Mia1 (43.1x) in zG as well as Ddah1 (zF/zG 16.2x), Cidea (15.5x), Frzb (9.5x), and Hsd11b2 (8.3x) in zF. The lists of transcripts obtained in the current study will be an invaluable tool for the elucidation of cellular mechanisms leading to zG and zF functional zonation.
Assessment of Epigenetic Contributions to Sexually-Dimorphic Kiss1 Expression in the Anteroventral Periventricular Nucleus of Mice
Authors: Semaan Sheila J. , Dhamija Sangeeta , Kim Joshua , Ku Eric C. , Kauffman Alexander S.
Published: February 2012

The Kiss1 gene, which encodes kisspeptin and is critical for reproduction, is sexually differentiated in the hypothalamic anteroventral periventricular (AVPV)/rostral periventricular (PeN) nuclei. Specifically, female rodents have higher AVPV/PeN Kiss1 expression than males, but how this Kiss1 sex difference is induced in early development is poorly understood. Here, we explored the contribution of epigenetic mechanisms to the establishment of the AVPV/PeN Kiss1 sex difference, focusing on histone deacetylation and DNA methylation. First, we utilized postnatal pharmacological blockade of histone deacetylation and analyzed Kiss1 expression in the AVPV/PeN. Postnatal disruption of histone deacetylase modestly increased AVPV Kiss1 cell number in both sexes but did not alter the Kiss1 sex difference. Next, we assessed whether the level of CpG methylation, which can influence transcription factor binding and gene expression, in the murine Kiss1 gene differs between males and females. We found significant sex differences in methylation at several CpG sites in the putative promoter and first intron of the Kiss1 gene in the AVPV/PeN, but not in the arcuate (which lacks adult Kiss1 sex differences), suggesting that differential methylation of the Kiss1 gene may influence sexually-dimorphic Kiss1 expression in the AVPV/PeN. Transgenic impairment of methyl CpG-binding protein-2 function did not eliminate the Kiss1 sex difference, indicating that other methylation factors are involved. Interestingly, CpG methylation in the AVPV/PeN was lower in males than females, suggesting that transcriptional repressors may contribute to the AVPV/PeN Kiss1 sex difference, a possibility supported by in silico identification of putative repressor binding sites near some of the sexually-dimorphic CpG.
Prolonged Exposure to Insulin Induces Mitochondrion-Derived Oxidative Stress through Increasing Mitochondrial Cholesterol Content in Hepatocytes
Authors: Mei Shuang , Gu Haihua , Yang Xuefeng , Guo Huailan , Liu Zhenqi , Cao Wenhong
Published: February 2012

We addressed the link between excessive exposure to insulin and mitochondrion-derived oxidative stress in this study and found that prolonged exposure to insulin increased mitochondrial cholesterol in cultured hepatocytes and in mice and stimulated production of reactive oxygen species (ROS) and decreased the reduced glutathione to glutathione disulfide ratio in cultured hepatocytes. Exposure of isolated hepatic mitochondria to cholesterol alone promoted ROS emission. The oxidative stress induced by the prolonged exposure to insulin was prevented by inhibition of cholesterol synthesis with simvastatin. We further found that prolonged exposure to insulin decreased mitochondrial membrane potential and the increased ROS production came from mitochondrial respiration complex I. Finally, we observed that prolonged exposure to insulin decreased mitochondrial membrane fluidity in a cholesterol synthesis-dependent manner. Together our results demonstrate that excess exposure to insulin causes mitochondrion-derived oxidative stress through cholesterol synthesis in hepatocytes.
Differential effects of Sec61α-, Sec62- and Sec63-depletion on transport of polypeptides into the endoplasmic reticulum of mammalian cells
Authors: Lang Sven , Benedix Julia , Fedeles Sorin V. , Schorr Stefan , Schirra Claudia , Schäuble Nico , Jalal Carolin , Greiner Markus , Haßdenteufel Sarah , Tatzelt Jörg , Kreutzer Birgit , Edelmann Ludwig , Krause Elmar , Rettig Jens , Somlo Stefan , Zimmermann Richard , Dudek Johanna
Published: February 2012

Co-translational transport of polypeptides into the endoplasmic reticulum (ER) involves the Sec61 channel and additional components such as the ER lumenal Hsp70 BiP and its membrane resident co-chaperone Sec63p in yeast. Here we investigated whether silencing the SEC61A1 gene in human cells affects co- and posttranslational transport of presecretory proteins into the ER and posttranslational membrane integration of tail-anchored proteins. While silencing the SEC61A1 gene in HeLa cells inhibited co- and posttranslational transport of signal peptide-containing precursor proteins into the ER of semi-permeabilized cells, silencing the SEC61A1 gene did not affect transport of various types of tail-anchored proteins. Furthermore, we demonstrate with a similar knock down approach a precursor-specific involvement of mammalian Sec63 in the initial phase of co-translational protein transport into the ER. In contrast, silencing the SEC62 gene inhibited only posttranslational transport of a signal-peptide containing precursor protein.
Plo1 phosphorylates Dam1 to promote chromosome bi-orientation in fission yeast
Authors: Buttrick Graham J. , Lancaster Theresa C. , Meadows John C. , Millar Jonathan B.A.
Published: February 2012

The fungal specific heterodecameric outer kinetochore DASH complex facilitates the interaction of kinetochores with spindle microtubules. In budding yeast, where kinetochores bind a single microtubule, the DASH complex is essential and phosphorylation of Dam1 by the Aurora kinase homologue, Ipl1, causes detachment of kinetochores from spindle microtubules. We demonstrate that in the distantly related fission yeast, where the DASH complex is not essential for viability and kinetochores bind multiple microtubules, Dam1 is instead phosphorylated on serine 143 by the Polo kinase homologue, Plo1, during prometaphase and metaphase. This phosphorylation site is conserved in most fungal Dam1 proteins, including budding yeast Dam1. We show Dam1 phosphorylation by Plo1 is dispensable for DASH assembly and chromosome retrieval but instead aids tension-dependent chromosome bi-orientation.
The IplA Ca++ Channel OfDictyostelium discoideumIs Necessary For Ca++, But Not cAMP Chemotaxis, And Plays A Fundamental Role In Natural Aggregation
Authors: Lusche Daniel F. , Wessels Deborah , Scherer Amanda , Daniels Karla , Kuhl Spencer , Soll David R.
Published: February 2012

During aggregation of Dictyostelium discoideum, nondissipating, symmetric,outwardly moving waves of cAMP direct cells towards aggregation centers. It has been assumed that the spatial and temporal characteristics of the front and back of each cAMP wave regulate both chemokinesis and chemotaxis. However, during the period preceding aggregation, cells acquire not only the capacity to chemotax in a spatial gradient of cAMP, but also in a spatial gradient of Ca++. The null mutant of the putative iplACa++ channel gene, iplA-, undergoes normal chemotaxis in spatial gradients of cAMP and normal chemokinetic responses to increasing temporal gradients of cAMP, both generated in vitro. However, iplA-cells lose the capacity to undergo chemotaxis in response to a spatial gradient of Ca++, suggesting that IplA is either the Ca++ chemotaxis receptor or an essential component of the Ca++ chemotaxis regulatory pathway. In response to natural chemotactic waves generated by wild type cells, the chemokinetic response of iplA- cells to the temporal dynamics of the cAMP waveis intact, but the capacity to reorient in the direction of the aggregation center at the onset of each waveis lost. These results suggest a model in which transient Ca++ gradients formed between cells at the onset of each natural cAMP wave augment reorientation towards the aggregation center. If this hypothesis proves correct, it will provide a more complex contextual framework for interpreting D. discoideum chemotaxis.
SUMO conjugation is required for the assembly ofDrosophilaSu(Hw) and Mod(mdg4) into insulator bodies that facilitate insulator complex formation
Authors: Golovnin Anton , Volkov Ilya , Georgiev Pavel
Published: February 2012

Chromatin insulators are special regulatory elements involved in modulation of enhancer–promoter interactions. The best studied insulators in Drosophila require Suppressor of Hairy Wing (Su(Hw)), Modifier of mdg4 (Mod(mdg4)) and centrosomal 190 kDa (CP190) proteins to be functional. These insulator proteins are co-localized in nuclear speckles named insulator bodies. Here, we demonstrate that post-translational modification of insulator proteins by Small Ubiquitin-like Modifier (SUMO) and intact CP190 protein is critical for insulator body formation. Inactivation of SUMO binding sites in Mod(mdg4)-67.2 leads to the inability for the mutant protein and Su(Hw) to be assembled into insulator bodies. In vivo functional tests show that a smaller amount of intact Mod(mdg4)-67.2, compared to the mutant protein, is required to restore the normal activity of the Su(Hw) insulator. However, high expression of mutant Mod(mdg4)-67.2 completely rescues the insulator activity, indicating that sumoylation is not necessary for enhancer blocking. These results suggest a model that insulator bodies function as a depot of sumoylated proteins that are involved in insulation and can facilitate insulator complex formation but are nonessential for insulator action.
Use of a bifurcated endovascular graft for treatment of endograft migration with major endoleak
Authors: Jim Jeffrey , Rubin Brian G , Sanchez Luis A
Published: February 2012

The purpose of this study was to describe the use of a bifurcated endovascular graft to treat endograft migration with major endoleaks. We present four patients who presented at a mean of 72.0 months after their initial endovascular abdominal aortic aneurysm repair. Three patients had type I endoleaks resulting from proximal attachment failure and graft migration. A fourth patient had separation of a proximal aortic cuff from a migrated main body device resulting in a type III endoleak. All were treated with a bifurcated Zenith (Cook Medical Incorporated, Bloomington, IN, USA) endovascular graft. There was 100% technical success with no perioperative complications. On follow-up, one patient died of unrelated causes at five months. The mean survival for the remaining three patients was 37 months. In conclusion, treatment with a bifurcated Zenith endograft has advantages over the use of an aortic cuff or aortouniiliac reconstruction. To perform this technique, there must be a sufficient distance between the proximal landing zone and the flow divider of the migrated endograft to allow for deployment of the Zenith device. While there remain limitations in its applicability, the use of a bifurcated endovascular graft is a viable alternative for endovascular salvage in treatment of endograft migration with major endoleaks.
Treatment for intermittent claudication and the effects on walking distance and quality of life
Authors: Kruidenier L M , Viechtbauer W , Nicolaï S P , Büller H , Prins M H , Teijink J A W
Published: February 2012

The objective of the study was to provide an overview of the most common treatments for intermittent claudication and to determine the effectiveness in improving walking distance and quality of life based on a combination of direct and indirect evidence. We included trials that compared: angioplasty, surgery, exercise therapy or no treatment for intermittent claudication. Outcome measurements were walking distance (maximum, pain-free) and quality of life (physical, mental). We used a network meta-analysis model for the combination of direct and indirect evidence. We included 42 studies, presenting 3106 participants. The network meta-analysis showed that supervised exercise therapy (Δ = 1.62, P < 0.01), angioplasty (Δ = 1.89, P < 0.01) and surgery (Δ = 2.72, P = 0.02) increased walking distance significantly more than no treatment. Furthermore, supervised exercise therapy (Δ = 0.60, P < 0.01), angioplasty (Δ = 0.91, P = 0.01) and surgery (Δ = 1.07, P < 0.01) increased physical quality of life more than no treatment. However, in the sensitivity analysis, only supervised exercise therapy had additional value over no symptomatic treatment (Δ = 0.66, P < 0.01). In conclusion, this network meta-analysis indicates that supervised exercise therapy is more effective in both increasing walking distance and physical quality of life, compared with no treatment. Angioplasty and surgery also increase walking distance, compared with no treatment, but results for physical quality of life are less convincing.
Percutaneous ultrasound-guided thermal ablation for intrahepatic cholangiocarcinoma
Authors: Xu H-X , Wang Y , Lu M-D , Liu L-N
Published: February 2012

Objective: The objective of this study was to evaluate the treatment efficacy and overall survival (OS) of percutaneous ultrasound-guided thermal ablation by means of microwave ablation or radiofrequency ablation for intrahepatic cholangiocarcinoma (ICC).

Methods: 18 patients with 25 ICC nodules underwent ultrasound-guided thermal ablation with curative intention. 8 patients were primary cases and 10 were recurrent cases after curative resection. The local treatment response, complications and survivals were analysed.

Results: Complete ablation was achieved in 23 (92.0%, 23/25) nodules (diameter, 0.7–4.3 cm; mean, 2.5±0.9 cm) and incomplete ablation was found in 2 (8.0%, 2/25) larger tumours (6.4 and 6.9 cm in diameter). No death associated with the treatment was found. The major complication rate was 5.5% (1/18). The follow-up periods ranged from 1.3 to 86.2 months (mean, 20.5±26.3 months; median, 8.7 months). OS rates for all patients at 6, 12, 36 and 60 months were 66.7%, 36.3%, 30.3% and 30.3%, respectively. By univariate analysis, the patient source (primary or recurrent case) was found to be a significant prognostic factor for OS rates (p=0.001). The patient source (p=0.001) and the number of nodules (p=0.038) were found to be significant prognostic factors for recurrence-free survival. OS rates for the primary ICC at 6, 12, 36 and 60 months were 87.5%, 75.0%, 62.5% and 62.5%, respectively.

Conclusion: Percutaneous ultrasound-guided thermal ablation is a safe and effective therapeutic technique for ICC. Acceptable survival can be achieved in primary ICCs, whereas the prognosis of recurrent ICCs is relatively poor.
Onyx 18 embolisation of dural arteriovenous fistula via arterial and venous pathways: preliminary experience and evaluation of the short-term outcomes
Authors: Long X-A , Karuna T , Zhang X , Luo B , Duan C-Z
Published: February 2012

Objective: This paper mainly focuses on our preliminary experience and short-term outcome evaluation of embolisation of non-cavernous dural arteriovenous fistulas (ncsDAVFs) and cavernous sinus dural arteriovenous fistulas (csDAVFs) using Onyx 18 (ev3, Plymouth, MN), and in combination with coils, via arterial and venous approaches, respectively.

Methods: Between August 2008 and March 2010, 21 DAVFs (11 ncsDAVFs and 10 csDAVFs; age range: 28–68 years; 12 females and 9 males) were undertaken. Borden classification showed Type III in 1 and Type II in 10 ncsDAVFs, and Type II in 4 and Type I in 6 csDAVFs. Onyx 18 was used in 11 ncsDAVFs (10 via single feeder and 1 via 2 feeders). Onyx 18 or in combination with coils was used in 10 csDAVFs (9 via the inferior petrosal sinus and 1 via the superior ophthalmic vein).

Results: Total occlusion in immediate angiography was achieved in 18 cases (85.7%; 10 ncsDAVFs and 8 csDAVFs), and near-total occlusion in 1 ncsDAVF and 2 csDAVFs. Onyx 18 was migrated into normal vasculature in two ncsDAVFs without any sequelae. One csDAVF had VI cranial nerve palsy post-operatively, which completely recovered 2 weeks post-embolisation. Follow-up angiography at 3–12 months showed complete occlusion in 20 cases (95.2%; 10 ncsDAVFs and 10 csDAVFs). One ncsDAVF (4.8%) recurred after 3 months and was successfully re-embolised.

Conclusion: Preliminary results achieved after embolising 11 ncsDAVFs and 10 csDAVFs using Onyx 18 and in combination with coils via arterial and venous pathways, respectively, appeared to be safe, feasible and effective, as 95.2% of cases were totally occluded without any clinical sequelae.
Sparseness of the trabecular pattern on dental radiographs: visual assessment compared with semi-automated measurements
Authors: Geraets W G M , Lindh C , Verheij H
Published: February 2012

Objective: In diagnostic imaging; human perception is the most prominent, yet least studied, source of error. A better understanding of image perception will help to improve diagnostic performance. This study focuses on the perception of coarseness of trabecular patterns on dental radiographs. Comparison of human vision with machine vision should yield knowledge on human perception.

Method: In a study on identifying osteoporotic patients, dental radiographs were made from 505 post-menopausal women aged 45–70 years. Intra-oral radiographs of the lower and upper jaws were made. Five observers graded the trabecular pattern as dense, sparse or mixed. The five gradings were combined into a single averaged observer score per jaw. The radiographs were scanned and a region of interest (ROI) was indicated on each. The ROIs were processed with image analysis software measuring 25 image features. Pearson correlation and multiple linear regression were used to compare the averaged observer score with the image features.

Results: 14 image features correlated significantly with the observer judgement for both jaws. The strongest correlation was found for the average grey value in the ROI. Other features, describing that osteoporotic patients have fewer but bigger marrow spaces than controls, correlated less with the sparseness of the trabecular pattern than a rather crude measure for structure such as the average grey value.

Conclusion: Human perception of the sparseness of trabecular patterns is based more on average grey values of the ROI than on geometric details within the ROI.
Effect of CT fluoroscopy-guided transpulmonary radiofrequency ablation of liver tumours on the lung
Authors: Iguchi T , Inoue D , Yabushita K , Sakaguchi K , Tatsukawa M , Sasaki H , Kanazawa S
Published: February 2012

Objective: We retrospectively evaluated the effect of transpulmonary radiofrequency ablation (RFA) of liver tumours on the lung.

Methods: 16 patients (10 males and 6 females; mean age, 65.2 years) with 16 liver tumours (mean diameter 1.5 cm) underwent transpulmonary RFA under CT fluoroscopic guidance. The tumours were either hepatocellular carcinoma (n=14) or liver metastasis (n=12). All 16 liver tumours were undetectable with ultrasonography. The pulmonary function values at 3 months after transpulmonary RFA were compared with baseline (i.e. values before RFA).

Results: In 8 of 16 sessions, minor pulmonary complications occurred, including small pneumothorax (n=8) and small pleural effusion (n=1). In two sessions, major pulmonary complications occurred, including pneumothorax requiring a chest tube (n=2). These chest tubes were removed at 4 and 6 days, and these patients were discharged 7 and 10 days after RFA, respectively, without any sequelae. The pulmonary function values we evaluated were forced expiratory volume in 1 s (FEV1.0) and vital capacity (VC). The mean values of FEV1.0 before and 3 months after RFA were 2.55 l and 2.59 l, respectively; the mean values of VC before and 3 months after RFA were 3.20 l and 3.27 l, respectively. These pulmonary values did not show any significant worsening (p=0.393 and 0.255 for FEV1.0 and VC, respectively).

Conclusion: There was no significant lung injury causing a fatal or intractable complication after transpulmonary RFA of liver tumours.
Recommendations for the use of PET and PET–CT for radiotherapy planning in research projects
Authors: Somer E J , Pike L C , Marsden P K
Published: February 2012

With the increasing use of positron emission tomography (PET) for disease staging, follow-up and therapy monitoring in a number of oncological indications there is growing interest in the use of PET and PET–CT for radiation treatment planning. In order to create a strong clinical evidence base for this, it is important to ensure that research data are clinically relevant and of a high quality. Therefore the National Cancer Research Institute PET Research Network make these recommendations to assist investigators in the development of radiotherapy clinical trials involving the use of PET and PET–CT. These recommendations provide an overview of the current literature in this rapidly evolving field, including standards for PET in clinical trials, disease staging, volume delineation, intensity modulated radiotherapy and PET-augmented planning techniques, and are targeted at a general audience. We conclude with specific recommendations for the use of PET in radiotherapy planning in research projects.