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11.
Javier Farinas Malea Kneen Megan Moore A.S. Verkman 《The Journal of general physiology》1997,110(3):283-296
A method was developed to measure the osmotic water permeability (Pf) of plasma membranes in cell layers and applied to cells and epithelia expressing molecular water channels. It was found that the integrated intensity of monochromatic light in a phase contrast or dark field microscope was dependent on relative cell volume. For cells of different size and shape (Sf9, MDCK, CHO, A549, tracheal epithelia, BHK), increased cell volume was associated with decreased signal intensity; generally the signal decreased 10–20% for a twofold increase in cell volume. A theory relating signal intensity to relative cell volume was developed based on spatial filtering and changes in optical path length associated with cell volume changes. Theory predictions were confirmed by signal measurements of cell layers bathed in solutions of various osmolarities and refractive indices. The excellent signal-to-noise ratio of the transmitted light detection permitted measurement of cell volume changes of <1%. The method was applied to characterize transfected cells and tissues that natively express water channels. Pf in control Chinese hamster ovary cells was low (0.0012 cm/s at 23°C) and increased more than fourfold upon stable transfection with aquaporins 1, 2, 4, or 5. Pf in apical and basolateral membranes in polarized epithelial cells grown on porous supports was measured. Pf
bl and Pf
ap were 0.0011 and 0.0024 cm/s (MDCK cells), and 0.0039 and 0.0052 cm/s (human tracheal cells) at 23°C. In intact toad urinary bladder, basolateral Pf was 0.036 cm/s and apical membrane Pf after vasopressin stimulation was 0.025 cm/s at 23°C. The results establish light microscopy with spatial filtering as a technically simple and quantitative method to measure water permeability in cell layers and provide the first measurement of the apical and basolateral membrane permeabilities of several important epithelial cell types. 相似文献
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Priscila da Silva Delabona Deise Juliana Lima Diogo Robl Sarita Cândida Rabelo Cristiane Sanchez Farinas José Geraldo da Cruz Pradella 《Journal of industrial microbiology & biotechnology》2016,43(5):617-626
The use of glycerol obtained as an intermediate of the biodiesel manufacturing process as carbon source for microbial growth is a potential alternative strategy for the production of enzymes and other high-value bioproducts. This work evaluates the production of cellulase enzymes using glycerol for high cell density growth of Trichoderma harzianum followed by induction with a cellulosic material. Firstly, the influence of the carbon source used in the pre-culture step was investigated in terms of total protein secretion and fungal morphology. Enzymatic productivity was then determined for cultivation strategies using different types and concentrations of carbon source, as well as different feeding procedures (batch and fed-batch). The best strategy for cellulase production was then further studied on a larger scale using a stirred tank bioreactor. The proposed strategy for cellulase production, using glycerol to achieve high cell density growth followed by induction with pretreated sugarcane bagasse, achieved enzymatic activities up to 2.27 ± 0.37 FPU/mL, 106.40 ± 8.87 IU/mL, and 9.04 ± 0.39 IU/mL of cellulase, xylanase, and β-glucosidase, respectively. These values were 2 times higher when compared to the control experiments using glucose instead of glycerol. This novel strategy proved to be a promising approach for improving cellulolytic enzymes production, and could potentially contribute to adding value to biomass within the biofuels sector. 相似文献
16.
Shim da J Nemeria NS Balakrishnan A Patel H Song J Wang J Jordan F Farinas ET 《Biochemistry》2011,50(35):7705-7709
The first component (E1o) of the Escherichia coli 2-oxoglutarate dehydrogenase complex (OGDHc) was engineered to accept substrates lacking the 5-carboxylate group by subjecting H260 and H298 to saturation mutagenesis. Apparently, H260 is required for substrate recognition, but H298 could be replaced with hydrophobic residues of similar molecular volume. To interrogate whether the second component would allow synthesis of acyl-coenzyme A derivatives, hybrid complexes consisting of recombinant components of OGDHc (o) and pyruvate dehydrogenase (p) enzymes were constructed, suggesting that a different component is the "gatekeeper" for specificity for these two multienzyme complexes in bacteria, E1p for pyruvate but E2o for 2-oxoglutarate. 相似文献
17.
M. N. Esperança F. M. Cunha M. O. Cerri T. C. Zangirolami C. S. Farinas A. C. Badino 《Bioprocess and biosystems engineering》2014,37(5):805-812
Sugarcane bagasse is a low-cost and abundant by-product generated by the bioethanol industry, and is a potential substrate for cellulolytic enzyme production. The aim of this work was to evaluate the effects of air flow rate (Q AIR), solids loading (%S), sugarcane bagasse type, and particle size on the gas hold-up (ε G) and volumetric oxygen transfer coefficient (k L a) in three different pneumatic bioreactors, using response surface methodology. Concentric tube airlift (CTA), split-cylinder airlift (SCA), and bubble column (BC) bioreactor types were tested. Q AIR and %S affected oxygen mass transfer positively and negatively, respectively, while sugarcane bagasse type and particle size (within the range studied) did not influence k L a. Using large particles of untreated sugarcane bagasse, the loop-type bioreactors (CTA and SCA) exhibited higher mass transfer, compared to the BC reactor. At higher %S, SCA presented a higher k L a value (0.0448 s?1) than CTA, and the best operational conditions in terms of oxygen mass transfer were achieved for %S < 10.0 g L?1 and Q AIR > 27.0 L min?1. These results demonstrated that pneumatic bioreactors can provide elevated oxygen transfer in the presence of vegetal biomass, making them an excellent option for use in three-phase systems for cellulolytic enzyme production by filamentous fungi. 相似文献
18.
Waka Yokoyama Hitoshi Kohsaka Kayoko Kaneko Matthew Walters Aiko Takayasu Shin Fukuda Chie Miyabe Yoshishige Miyabe Paul E Love Nobuhiro Nakamoto Takanori Kanai Kaori Watanabe-Imai Trevor T Charvat Mark ET Penfold Juan Jaen Thomas J Schall Masayoshi Harigai Nobuyuki Miyasaka Toshihiro Nanki 《Arthritis research & therapy》2014,16(5)
Introduction
Biological drugs are effective in patients with rheumatoid arthritis (RA), but increase severe infections. The CC chemokine receptor (CCR) 9 antagonist was effective for Crohn’s disease without critical adverse effects including infections in clinical trials. The present study was carried out to explore the pathogenic roles of chemokine (C-C motif) ligand (CCL) 25 and its receptor, CCR9, in autoimmune arthritis and to study if the CCR9 antagonist could be a new treatment for RA.Methods
CCL25 and CCR9 expression was examined with immunohistochemistry and Western blotting. Concentration of interleukin (IL)-6, matrix metalloproteinase (MMP)-3 and tumor necrosis factor (TNF)-α was measured with enzyme-linked immunosorbent assays. Effects of abrogating CCR9 on collagen-induced arthritis (CIA) was evaluated using CCR9-deficient mice or the CCR9 antagonist, CCX8037. Fluorescence labeled-CD11b+ splenocytes from CIA mice were transferred to recipient CIA mice and those infiltrating into the synovial tissues of the recipient mice were counted.Results
CCL25 and CCR9 proteins were found in the RA synovial tissues. CCR9 was expressed on macrophages, fibroblast-like synoviocytes (FLS) and dendritic cells in the synovial tissues. Stimulation with CCL25 increased IL-6 and MMP-3 production from RA FLS, and IL-6 and TNF-α production from peripheral blood monocytes. CIA was suppressed in CCR9-deficient mice. CCX8037 also inhibited CIA and the migration of transferred CD11b+ splenocytes into the synovial tissues.Conclusions
The interaction between CCL25 and CCR9 may play important roles in cell infiltration into the RA synovial tissues and inflammatory mediator production. Blocking CCL25 or CCR9 may represent a novel safe therapy for RA. 相似文献19.
Rosangela D. P. B. Pirota Priscila S. Delabona Cristiane S. Farinas 《Bioenergy Research》2014,7(2):744-752
A novel simplified configuration is proposed for the conversion of biomass to ethanol using whole medium enzymatic cocktails (WM) and enzymatic extracts (EE) from different filamentous fungi (Trichoderma reesei, Aspergillus niger, and Aspergillus oryzae) cultivated under solid-state fermentation (SSF) for the hydrolysis of steam-exploded sugarcane bagasse (SESB). The hydrolyzed material derived from the saccharification of SESB using the combinations A. niger WM + T. reesei EE, A. oryzae WM + A. niger EE, and A. niger EE + T. reesei WM resulted in the best biomass conversion yields (66, 65, and 64 % of the theoretical reducing sugar yields, respectively). The best ethanol production (84 % of the theoretical yield) was obtained using the material hydrolyzed by a combination of A. oryzae WM + A. niger EE. The enzymatic conversion of SESB using on-site produced enzymes from the whole SSF cultivation medium, followed by an ethanol production step, is a potential configuration for the biomass to ethanol conversion process. This novel simplified configuration would enable the use of a single reactor system, avoiding the need for additional separation steps. 相似文献
20.
PIERRE MOISSETTE ET JEAN-PAUL SAINT MARTIN 《Lethaia: An International Journal of Palaeontology and Stratigraphy》1995,28(3):271-283
Moissette, P. & Saint Martin, J:P. 1995 11 30 Bryozoaires des milieux récifaux miocenes du sillon sud-rifain au Maroc. [Bryozoans from Miocene reef environments in the South-Rifian corridor of Morocco. In late Miocene coral reefs from the Fes area (Northern Morocco), bryozoans are the best-represented group. An inventory of the bryozoan fauna (59 species) was taken on two reefs with different organization and palaeogeographical situation in a late Miocene seaway between the Atlantic and the Mediterranean. A study of the bryozoans and their zoarial forms allows an improved reconstruction of reefal environments and available habitats. Membraniporiforrn (encrusting) colonies largely predominate, whereas celleporiforms (nodular colonies) are more rare. Rigid erect (vinculariiform, adeoniform, and reteporiform) and rigid articulated zoarial types (cellariiform and catenicelliform) are fairly well represented. At depths of about 10 m and on the reef front encrusting bryozoans were well developed in cryptic habitats on living coral substrates. Erect species lived in cavities provided by the coral framework, and plants offered flexible substrates for the attachment of epiphytal forms. □Bryozoa, coral reefs, Morocco, Messininn, pulueoecology. Dans les récifs coralliens du Miocène supérieur de la région de Fes (Maroc septentrional), les bryozoaires constituent, parrni les organismes conservés, le groupe le mieux représenté. Un recensement de la fame de bryozoaires (59 espèces) a été effectué dans deux récifs d'organisation et de situation paléogéographique différentes, dans un secteur de communication entre Atlantique et Méditerranée au MiocéFne supérieur. L'étude des bryozoaires et de leurs formes zoariales permet de proposer une meilleure reconstitution de I'environnement récifal et des habitats disponibles. Les colonies membraniporiformes (encrobtantes) predominent largement, alors que les cellepori-formes (colonies noduleuses) sont beaucoup plus rares. Les types zoariaux érigés rigides (vincula-riiforme, adéoniforme et rétéporiforme) et érigés articulés (cellariiforme et catenicelliforme) sont assez bien représentés. A des profondeurs denviron 10 m et dans la partie antérieure des constructions récifales, les bryozoaires encroûtants étaient bien développés dans des habitats cryptiques sur substrats coralliens vivants. Des formes dressées vivaient dans les cavités ménagées par le bâti corallien et des supports végétaux permettaient également la fixation de formes épiphytes. □Bryozouires, récifs coralliens, Muroc, Messinien, puléoécologie. 相似文献