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Article No. Title
UH1081 Comprehensive phospholipid and sphingomyelin profiling of different brain regions in mouse model of anxiety disorder using online two-dimensional (HILIC/RP)-LC/MS method
  Róbert Berkecz, Ferenc Tömösi, Tímea Körmöczi, Viktor Szegedi, János Horváth, Tamás Janáky
PDF icon A novel online system including two-dimensional liquid chromatography coupled to high-resolution mass spectrometry (2D-LC/MS) was developed and applied for comprehensive phospholipid (PL) and sphingomyelin (SM) profiling of dorsal hippocampus (DHPC), ventral (VHPC) and prefrontal cortex (PFC) brain regions in a mouse model of anxiety disorder. In the first dimension, lipid classes were distinguished by hydrophilic interaction liquid chromatography (HILIC), while the second dimensional separation of individual PL and SM species was achieved by reversed-phase (RP) chromatography. For the enrichment of lipid species in diluted HILIC effluent, two RP trapping columns were used separately. The developed fully-automated 2D method allowed the quantitative analysis of over 150 endogenous PL and SM species in mouse brain regions within 40 min. The developed method was applied in a pilot study, which aimed to find alteration of PL and SM composition in a mouse model of anxiety disorder. In the case of 37 PL and SM species, significant differences were observed between high anxiety-related behavior (AX) and low anxiety-related behavior (nAX) mice. In mice having elevated anxiety, the most typical trend was the downregulation of PL species, in particular, in VHPC.
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UH1082 Mutations of KCNJ10 Together with Mutations of SLC26A4 Cause Digenic Nonsyndromic Hearing Loss Associated with Enlarged Vestibular Aqueduct Syndrome
  Tao Yang, Jose G. Gurrola II, Hao Wu, Sui M. Chiu, Philine Wangemann, Peter M. Snyder, and Richard J.H. Smith
PDF icon Mutations in SLC26A4 cause nonsyndromic hearing loss associated with an enlarged vestibular aqueduct (EVA, also known as DFNB4) and Pendred syndrome (PS), the most common type of autosomal-recessive syndromic deafness. In many patients with an EVA/PS phenotype, mutation screening of SLC26A4 fails to identify two disease-causing allele variants. That a sizable fraction of patients carry only one SLC26A4 mutation suggests that EVA/PS is a complex disease involving other genetic factors. Here, we show that mutations in the inwardly rectifying Kþ channel gene KCNJ10 are associated with nonsyndromic hearing loss in carriers of SLC26A4 mutations with an EVA/PS pheno- type. In probands from two families, we identified double heterozygosity in affected individuals. These persons carried single mutations in both SLC26A4 and KCNJ10. The identified SLC26A4 mutations have been previously implicated in EVA/PS, and the KCNJ10 mutations reduce Kþ conductance activity, which is critical for generating and maintaining the endocochlear potential. In addition, we show that hap- loinsufficiency of Slc26a4 in the Slc26a4þ/􏰀 mouse mutant results in reduced protein expression of Kcnj10 in the stria vascularis of the inner ear. Our results link KCNJ10 mutations with EVA/PS and provide further support for the model of EVA/PS as a multigenic complex disease.
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UH1083 Linamarase Enzyme from Lactobacillus delbrueckii NRRL B-763: Purification and Some Properties of a β-Glucosidase
  Ogbonnaya Nwokoro and Florence Onyebuchi Anya
PDF icon Some biochemical properties and purification of linamarase enzyme from Lactobacillus delbrueckii NRRL B-763 were studied. The crude enzyme was used to detoxify cassava flour cyanide and samples of 150 μm particle size treated with the crude enzyme showed a reduction from 2.1 mg HCN/10g sample to 0.11 mg HCN/10 g sample after 20 h (95% reduction). Untreated control samples of 0.5 mm particle size showed a reduction from 2.1mg HCN/10 g sample to 1.98 mg HCN/10 g sample after 40 h (5.7% reduction). The enzyme was purified 33 fold with a 40% yield through a series of four steps namely, ammonium sulphate precipitation, acetone precipitation, ion exchange chromatography and gel filteration chromatograrphy using Sephadex G-200. The purified enzyme showed maximum activity at pH 4.5. The enzyme showed 100% stability at the pH range of 5.0 and 6.0. Maximum activity of the enzyme was observed at a temperature of 50 oC and maximum stability at a temperature range of 40 and 50 oC. The approximate enzyme molecular weight was estimated to be 56 kDa by Sephadex G-200 gel filteration chromatography. The linamarase enzyme could be adapted for improved degradation of cassava cyanide and other biotechnological applications.
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UH1084 Studies on the production of glucose isomerase by Bacillus licheniformis
  Ogbonnaya Nwokoro
PDF icon This work reports the effects of some culture conditions on the production of glucose isomerase by Bacillus licheniformis.The bacterium was selected based on the release of 3.62 mg/mL fructose from the fermentation of glucose. Enzyme was produced using a variety of carbon substrates but the highest enzyme activity was detected in a medium containing 0.5% xylose and 1% glycerol (specifi c activity = 6.88 U/mg protein). Media containing only xylose or glucose gave lower enzyme productivies (specifi c activities= 4.60 and 2.35 U/mg protein respectively).
The effects of nitrogen substrates on glucose isomerase production showed that yeast extract supported maximum enzyme activity (specifi c activity = 5.24 U/mg protein). Lowest enzyme activity was observed with sodium trioxonitrate (specifi c activity = 2.44 U/mg protein). In general, organic nitrogen substrates supported higher enzyme productivity than inorganic nitrogen substrates. Best enzyme activity was observed in the presence of Mg2+ (specifi c activity = 6.85 U/mg protein) while Hg2+ was inhibitory (specifi c activity = 1.02 U/mg protein). The optimum pH for best enzyme activity was 6.0 while optimum temperature for enzyme production was 50oC.
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UH1085 Genotoxicity Studies of Titanium Dioxide Nanoparticles (TiO2NPs) in the Brain of Mice
  Hanan R. H. Mohamed and Nahed A. Hussien
PDF icon Titaniumdioxide nanoparticles (TiO2NPs) are excessively used and represent one of the top five most commonly used nanoparticles worldwide. Recently, various studies referred to their toxic potential on various organs using different treatment route. Male Swiss Webster mice were orally administrated TiO2NPs (500mg/kg b.w.) daily for five consecutive days and then animals were sacrificed at 24 h, 7 days, or 14 days after the last treatment.The present results report that exposure to TiO2NPs produces mild to moderate changes in the cytoarchitecture of brain tissue in a time dependent manner. Moreover, Comet assay revealed the apoptotic DNA fragmentation, while PCR-SSCP pattern and direct sequencing showed pointmutation of Presenilin 1 gene at exon 5, gene linked to inherited forms of the Alzheimer’s disease. Therefore, from these findings, the present study concluded that TiO2NPs is genotoxic and mutagenic to brain tissue which in turn might lead to Alzheimer’s disease incidence.
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UH1086 The Na+/H+ exchanger contributes to increased smooth muscle proliferation and migration in a rat model of pulmonary arterial hypertension
  John C. Huetsch, Haiyang Jiang, Carolina Larrain & Larissa A. Shimoda
PDF icon Increased muscularity of small pulmonary vessels, involving enhanced proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs), is a key component of the vascular remodeling underlying the development of pulmonary hypertension (PH). Stimuli such as growth factors and hypoxia induce PASMC alkalinization, proliferation, and migration through upregulation of the Na+/H+ exchanger (NHE), inhibition of which prevents the development of hypoxia-induced vascular remodeling and PH. We wanted to explore whether NHE was also necessary for pathologic PASMC proliferation and migration in a model of pulmonary arterial hypertension (PAH), a severe form of PH not associated with persistent hypoxia. PASMCs were isolated from rats exposed to SU5416-hypoxia (SuHx) followed by return to normoxia and from vehicle controls. We measured resting intracellular pH (pHi) and NHE activity using the pH-sensitive fluorescent dye BCECF-AM. PASMC proliferation and migration were assessed using BrdU incorporation and transwell filters, respectively. NHE activity was increased in SuHx PASMCs, although resting pHi was unchanged. SuHx PASMCs also exhibited increased proliferation and migration relative to controls, which was attenuated in the setting of pharmacologic inhibition of NHE. Our findings suggest that increased NHE activity contributes to pathologic PASMC function in the SuHx model of PAH, although this effect does not appear to be mediated by global changes in pHi homeostasis.
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UH1087 Attenuation of Nano-Tio2 Induced Genotoxicity, Mutagenicity and Apoptosis by Chlorophyllin in
Mice Cardiac Cells
  Hanan Ramadan Hamad Mohamed
PDF icon Despite uses of titanium dioxide (TiO2) particles in various consumer and medical products therapy increasing human daily exposure, little studies were conducted on its cardiotoxicity. Thus, this study was designed to investigate the possible modulation of nano- TiO2 induced genotoxicity, mutagenicity and apoptosis by chlorophyllin (CHL) coadministration in mice cardiac cells. Mice were injected into the abdominal cavity with TiO2 (500, 1000 and 2000 mg/kg b.w) suspended either in deionized dist. water or in CHL (40 mg/kg b.w) solution for five consecutive days and scarified 24 hour after the last injections. CHL co-administration resulting in significant reductions in tail length, %DNA in tail and tail moment that was highly elevated in nano-TiO2 reated groups in a dose dependent manner, also the appearance of both apoptotic fragmentized Laddered and smeared DNA on agarose gel was returned to the normal unfragmentized appearance. The observed dose dependent high incidence of mutations inductions in p53 exons (5-8) and myocardial cells infiltrations by inflammatory cells, hemorrhage and congested blood vessels by nano-titanium was attenuated by CHL co-administration. Finally, CHL improved the antioxidant defense system indicated by significant reduction in malondialdiahyde level and increases in reduced glutathione level and glutathione peroxidase activity that were disrupted in nano-titanium treated groups. In conclusion: nano-TiO2 particles induced genotoxicity, mutagenicity and apoptosis were attenuated by CHL co-administration in mice cardiac cells. It is recommended for further studies to detect the CHL dose that completely normalize nano-titanium induced cardiotoxicity.
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UH1088 Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications
  Tiago M. Fortunato1, Cristina Beltrami, Costanza Emanueli, Paul A. De Bank1 & Giordano Pula
PDF icon Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures. Additional advantages of our approach over existing extracellular matrices are the absence of any animal product in the composition hPLG and the possibility of obtaining hPLG from patients to generate homologous scaffolds for re-implantation. This discovery has the potential to accelerate the development of regenerative medicine applications based on implantation of microvascular networks expanded ex vivo or the generation of fully vascularisedorgans.
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UH1089 Construction and Preliminary Screening of Metagenomic Library from Lonar Crater Lake, India Ms. Priya Borekar
  Ms. Priya Borekar
PDF icon Lonar Lake is a eutrophic, saline soda lake with permanently anoxic deep water. Saline water lake microbes are a large and diverse group, which are exposed to a wide variety of pressure, temperature, salinity, nutrient availability and other environmental conditions. Soil metagenomics, which comprises isolation of soil DNA and the production and screening of clone libraries, can provide a cultivation-independent assessment of the largely untapped genetic reservoir of soil microbial communities. Metagenomics as a new field of research has been developed over the past decade to analyzed collective genomes of an assemblage of organisms, of the non-cultured microbes with the goal to better understand global microbial ecology on the one side and on the other side has been successfully applied to isolate novel biocatalysts from the uncultured microbiota in the environment. Recent progress in molecular microbial ecology has been applied to diverse problems in microbiology and has yielded insight into the physiology of uncultured organisms to access the potentially useful enzymes and secondary metabolites they produce. They provide a huge potential source of novel enzymes with unique properties that may be useful in biotechnological industry and further development of high throughput technologies for accelerating the pace of discovery in microbial ecology.
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PDF icon Six cyanobacterial strains isolated from different locations in Egypt were screened for the presence of restriction enzyme(s) where the extraction of restriction enzymes was performed using two-buffer system. Species of Phormidium autumnale, Cyanothece aeruginosa and Spirulina platensis were found to possess restriction activity that linearized plasmid pUC18 DNA. To identify the protein fragments of restriction activity, a DEAE cellulose column fractionation was performed and elution was carried out by gradient concentrations of NaCl. Fractions isolated with concentration of 0.1 and 0.4 M NaCl were found to possess restriction activity. Biotechnological potential of those fractions was tested using genomic DNAs from two different plants. Both the genomic DNAs from Lactuca sativa and Solanum tuberosum were successfully cut thereby confirming their biotechnological potential.
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Ultrasonic Homogenizers
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Our Ultrasonic Homogenizer product line is being expanded to include thermo-conductive tube pods. KoldPods are available for 1.5ml Micro Tubes, 15ml and 50ml Conical tubes, and will decrease thermal variability with conventional temperature sources.

Ultrasonic Homogenizers are also referred to as Sonicators, Cell Disruptors, Probe Sonicators, Sonifiers®, and Sonic Dismembrators

Sonifier is a registered trademark of Branson Ultrasonics Corporation

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