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At least 577 records · Page 32Linked to original sources
PubMed1992

Downregulation of Newcastle disease virus (NDV)-dependent IFN-alpha/beta production in macrophages by IFN-induced gene products of the locus If-1.

We have found that in cultured mouse bone-marrow-derived macrophages (BMDM), endogenous IFN-beta specifically regulates Newcastle disease virus (NDV) induced interferon (IFN)-alpha/beta synthesis, possibly by influencing the activity of genes within the regulatory locus If-1. Comparison of anti-IFN-beta-treated BMDM from C57BL/6 mice and the congenic line B.6C-H28c carrying the high (h) or low (l) producer allele of If-1, respectively, revealed a much stronger response of the If-1l allele to exogenous IFN-alpha treatment. Twenty IU rIFN-alpha 4 were sufficient to induce nearly complete suppression of NDV-induced IFN-alpha and IFN-beta production in BMDM from B6.C-H28c mice, but had no effect on the IFN-alpha/beta response induced by Sendai virus, another member of the paramyxovirus group. Simultaneous treatment of BMDM with cycloheximide inhibited the suppressive effect of rIFN-alpha 4, indicating that IFN induced the expression of one or several new proteins encoded by gene(s) within the If-1l locus which are responsible for the NDV-specific downregulation of IFN-alpha/beta production. A time course analysis indicated that the suppressive activity of IFN-induced If-1l gene products took 12 h to develop. It was preceded by an opposite priming effect, leading to enhancement of the early IFN-alpha/beta response to NDV measured 5 h after infection. This priming effect in BMDM was, however, only visible during an 8-h period of IFN-alpha treatment, whereas in the continued presence of IFN for 12 h or longer, priming was superimposed by the inhibitory action of the If-1l gene products.(ABSTRACT TRUNCATED AT 250 WORDS)

Alleles↗
PubMed2005

Glucoamylase production by solid-state fermentation using rice flake manufacturing waste products as substrate.

Glucoamylase production has been investigated by solid-state fermentation of agro-industrial wastes generated during the processing of paddy to rice flakes (categorized as coarse, medium and fine waste), along with wheat bran and rice powder by a local soil isolate Aspergillus sp. HA-2. Highest enzyme production was obtained with wheat bran (264 +/- 0.64 U/gds) followed by coarse waste (211.5 +/- 1.44 U/gds) and medium waste (192.1 +/- 1.15 U/gds) using 10(6) spores/ml as inoculum at 28 +/- 2 degrees C, pH 5. A combination of wheat bran and coarse waste (1:1) gave enzyme yield as compared to wheat bran alone. Media supplementation with carbon source (0.04 g/gds) as sucrose in wheat bran and glucose in coarse and medium waste increased enzyme production to 271.2 +/- 0.92, 220.2 +/- 0.75 and 208.2 +/- 1.99 U/gds respectively. Organic nitrogen supplementation (yeast extract and peptone, 0.02 g/gds) showed a higher enzyme production compared to inorganic source. Optimum enzyme activity was observed at 55 degrees C, pH 5. Enzyme activity was enhanced in the presence of calcium whereas presence of EDTA gave reverse effect.

Aspergillus↗
PubMed2005

Differences in product information of biopharmaceuticals in the EU and the USA: implications for product development.

The Summary of Product Characteristics (SPC) approved by the European Medicines Agency (EMEA) and the Package Insert (PI) approved by the Food and Drug Administration (FDA) were examined for 32 biopharmaceutical products. The aim was to identify differences in the product information since such information may have an impact on the planning of global clinical development programmes. The EU SPC contained more detailed instructions to the prescriber, including the positioning of the product with regard to the stage of the disease and to other therapies. The approach to safety information, notably to contraindications and warnings was more conservative in the EU SPC. The conservative approach in the EU may reflect the central position of the SPC in risk management of new pharmaceuticals. A typical feature of the US PI was the detailed description of the efficacy and safety result of the pivotal clinical trials.

Drug Design↗
PubMed2006

Adaptors discriminate mast-cell cytokine production from eicosanoid production and degranulation.

The Fc epsilonRI-dependent activation of nuclear factor (NF)kappaB is key for mast-cell cytokine production. The CARMA1-Bcl10-Malt1 adaptor complex regulates NFkappaB activation by antigen receptors in lymphocytes. A recent study reveals that the Bcl10-Malt1 complex promotes mast-cell interleukin-6 and tumor necrosis factor production, independent of degranulation, eicosanoid secretion or survival. The new findings place this complex at the forefront in discriminating between signals required for cytokine production and those required for mast-cell degranulation and eicosanoid production.

Adaptor Proteins, Signal Transducing↗
PubMed2004

Scale-up of virus-like particles production: effects of sparging, agitation and bioreactor scale on cell growth, infection kinetics and productivity.

The baculovirus-insect cells expression system was used for the production of self-forming Porcine parvovirus (PPV) like particles (virus-like particles, VLPs) in serum-free medium. At 2l bioreactor scale an efficient production was achieved by infecting the culture at a concentration of 1.5 x 10(6)cells/ml using a low multiplicity of infection of 0.05 pfu per cell. In a continuous bioreactor, it was shown that the uninfected insect cells were not sensitive to local shear stress values up to 2.25 N/m2 at high Reynolds numbers (1.5 x 10(4)) in sparging conditions. Uninfected insect cells can be grown at scaled-up bioreactor at high agitation and sparging rates as long as vortex formation is avoided and bubble entrapment is minimized. An efficient process scale-up to 25 l bioreactor was made using constant shear stress criteria for scale-up. The kinetics of baculovirus infection at low multiplicity of infection, either at different cell concentration or at different scales, are very reproducible, despite the different turbulence conditions present in the bioreactor milieu. The results suggest that the infection kinetics is controlled by the rate of baculovirus-cell receptor attachment and is independent of the bioreactor hydrodynamic conditions. Furthermore, the achieved specific and volumetric productivities were higher at the 25 l scale when compared to the smaller scale bioreactor. Different rates of cell lysis after infection were observed and seem to fully explain both the shift in optimal harvest time and the increase in cell specific productivity. The results emphasize the importance of integrated strategies and engineering concepts in process development at bioreactor stage with the baculovirus insect cell system.

Animals↗
PubMed2005

Triple layered rotavirus VLP production: kinetics of vector replication, mRNA stability and recombinant protein production.

Rotavirus infection causes diarrhoeal disease in infants, killing more than half million children each year. Virus-like particles (VLP) seem to be excellent vaccine candidates, since they are cheaper to produce than attenuated viral vaccines and safer, as they do not contain genetic material. The present work focus on a triple layered particle composed by three rotavirus structural proteins: VP2, VP6 and VP7, produced in an insect cell/baculovirus expressing system. Two strategies were evaluated for 2/6/7 VLP production: co-infection with three monocistronic baculovirus vectors or single-infection with a tricistronic multi-gene baculovirus vector; these strategies were followed at different levels: baculovirus DNA replication kinetics, mRNA stability, protein production and VLP formation. This study highlights some of the reasons why the tricistronic baculovirus strategy is more efficient for production of triple layered rotavirus 2/6/7 VLP than monocistronic co-infection, in particular: (i) the tricistronic vector presents higher DNA replication rates than the monocistronic vectors, (ii) the mRNA stability is invariant for all mRNAs corresponding to VP2, VP6 and VP7 and (iii) the tricistronic baculovirus strategy produces an excess of VP7 over VP6 when compared to the VP7/VP6 stoichiometric ratio in the native rotavirus. Although the co-infection strategy leads to protein production akin to the rotavirus VP7/VP6 stoichiometric ratio, the tricistronic vector strategy yields higher amounts of rotavirus-like particles.

Animals↗
PubMed2006

Characterization of a spliced exon product of herpes simplex type-1 latency-associated transcript in productively infected cells.

The latency-associated transcripts (LATs) of herpes simplex virus type-1 (HSV-1) are the only viral RNAs accumulating during latent infections in the sensory ganglia of the peripheral nervous system. The major form of LAT that accumulates in latently infected neurons is a 2 kb intron, spliced from a much less abundant 8.3 primary transcript. The spliced exon mRNA has been hard to detect. However, in this study, we have examined the spliced exon RNA in productively infected cells using ribonuclease protection (RPA), and quantitative RT-PCR (q-PCR) assays. We were able to detect the LAT exon RNA in productively infected SY5Y cells (a human neuronal cell line). The level of the LAT exon RNA was found to be approximately 5% that of the 2 kb intron RNA and thus is likely to be relatively unstable. Quantitative RT-PCR (q-PCR) assays were used to examine the LAT exon RNA and its properties. They confirmed that the LAT exon mRNA is present at a very low level in productively infected cells, compared to the levels of other viral transcripts. Furthermore, experiments showed that the LAT exon mRNA is expressed as a true late gene, and appears to be polyadenylated. In SY5Y cells, in contrast to most late viral transcripts, the LAT exon RNA was found to be mainly nuclear localized during the late stage of a productive infection. Interestingly, more LAT exon RNA was found in the cytoplasm in differentiated compared to undifferentiated SY5Y cells, suggesting the nucleocytoplasmic distribution of the LAT exon RNA and its related function may be influenced by the differentiation state of cells.

Cell Differentiation↗
PubMed2006

Natural sweetening of food products by engineering Lactococcus lactis for glucose production.

We show that sweetening of food products by natural fermentation can be achieved by a combined metabolic engineering and transcriptome analysis approach. A Lactococcus lactis ssp. cremoris strain was constructed in which glucose metabolism was completely disrupted by deletion of the genes coding for glucokinase (glk), EII(man/glc) (ptnABCD), and the newly discovered glucose-PTS EII(cel) (ptcBAC). After introducing the lactose metabolic genes, the deletion strain could solely ferment the galactose moiety of lactose, while the glucose moiety accumulated extracellularly. Additionally, less lactose remained in the medium after fermentation. The resulting strain can be used for in situ production of glucose, circumventing the need to add sweeteners as additional ingredients to dairy products. Moreover, the enhanced removal of lactose achieved by this strain could be very useful in the manufacture of products for lactose intolerant individuals.

Escherichia coli Proteins↗
PubMed1988

The in vitro activation of cyclic AMP production by either forskolin or isoproterenol in the Syrian hamster pineal during the day is not accompanied by an increase in melatonin production.

The role of cyclic AMP in the regulation of melatonin production was investigated in cultured Syrian hamster pineal glands. Forskolin markedly increased cyclic AMP production in pineal glands collected either late in the light period or in the dark period. The effect of forskolin was synergistically enhanced by 3-isobutylmethylxanthine, a phosphodiesterase inhibitor; however, increase in cyclic AMP after isoproterenol was only apparent in the presence of 3-isobutylmethylxanthine. Since beta-adrenergic agonists are able to stimulate melatonin production late in the dark period only, these data suggest that, in the hamster pineal gland, there may be intracellular mechanisms in addition to a cyclic AMP increase required for induction of melatonin production by beta-adrenergic agonists.

1-Methyl-3-isobutylxanthine↗
PubMed2002

Dialysis-based bioreactor systems for the production of monoclonal antibodies--alternatives to ascites production in mice.

Two commercially available bioreactor systems, CELLine and miniPERM, were evaluated for their ability to support the production of monoclonal antibody (mAb) from a variety of murine hybridoma cell lines. Production and purity of mAbs were compared between the two systems and with mouse ascites tumour fluid generation. The quality and purity of the mAb generated by each method was analysed on SDS-PAGE gels and the antibody immunoreactivity in each case was quantified by indirect ELISA tests. The relative benefits of conventional growth medium (Dulbecco's modified Eagle's media, DMEM) and serum-free medium (hybridoma serum-free media, H-SFM) using the miniPERM system were also analysed, in terms of the amount of antibody produced, cell concentration and specific antibody titre. In all cases, the CELLine units tested gave higher protein concentrations compared to the miniPERM system under the same conditions (means and 95% confidence limits are 4.2+/-0.8 and 2.1+/-0.8 mg/ml, respectively), yet the miniPERM system yielded greater total amounts over a similar culture period (428.7+/-243.3 mg compared to 183.3+/-100.9 mg in the CL-350 CELLine unit). When defined by specific ELISA titre, both bioreactor systems yielded mAb levels that compared favourably with those derived from ascites. In addition, SDS-PAGE analysis indicated that the bioreactor antibody product was relatively free of contaminating protein, whereas ascites tumour fluid preparations displayed significant levels of extraneous protein. This study has shown that both bioreactor systems are acceptable in vitro alternatives to the in vivo production of mAbs in mice.

Animals↗
PubMed2001

Hydrogen production in anaerobic reactors during shock loads--influence of formate production and H2 kinetics.

In this article the role of hydrogen as a process monitoring tool in methanogenic systems was studied by considering the influence of several key system parameters. Hydrogen production was found to be influenced mainly by the inocula's source pH, and varied only slightly with external pH and HCO3- levels. When an inoculum adapted to above neutral conditions (pH > 7) was shocked, reducing equivalents were selectively channelled through formate, while high hydrogen production was noticed with acidically (pH < 6.5) adapted inocula. The results also revealed that the production of hydrogen or formate during shock loads was not strongly associated with microbial morphology (granules or flocs) as high electron fluxes were possible through either during acidogenesis. Shock load experiments in continuous reactors revealed that neither hydrogen nor formate accumulated to any significant degree, nevertheless digester recovery took a long time due to the slow kinetics of volatile fatty acid degradation. Selective formate production under neutral pH environments, coupled with high hydrogenotrophic activity, was found to be responsible for the dampened hydrogen response during the early phases of gradually shocked systems (step change). Based on these results it appears that the role of hydrogen as a process monitoring tool has been overemphasised in the literature.

Bioreactors↗
PubMed1996

Immunotherapy suppresses the production of monocyte chemotactic and activating factor and augments the production of IL-8 in children with asthma.

BACKGROUND: Histamine-releasing factor consists of a group of cytokines that can cause basophils or mast cells to release histamine. However, the composition of histamine-releasing factor remains undefined. OBJECTIVE: This study was done to measure the concentrations, in plasma and mononuclear cell culture supernatants from children with asthma, of chemokines that are known to contribute to histamine-releasing factor activity. RESULTS: Plasma and mononuclear cell culture supernatants were obtained from 25 children newly diagnosed with asthma, 25 good responders to immunotherapy, 23 poor responders, 25 patients with acute attacks, and 13 normal subjects. All the patient groups produced, spontaneously and after stimulation with phytohemagglutinin and mite allergen, greater amounts of monocyte chemotactic and activating factor, macrophage inflammatory protein-1 alpha, and RANTES (beta chemokines) and IL-8 and growth-related gene alpha (alpha chemokines) than did normal subjects. Successful immunotherapy resulted in decreased production, especially the spontaneous type, of beta chemokines that cause histamine release and in increased production of a chemokines that inhibit histamine release. CONCLUSION: Abnormal chemokine production may contribute to the pathogenesis of bronchial asthma, and restoration of normal chemokine production may be used to explain, in part, the clinical efficacy of immunotherapy.

Adjuvants, Immunologic↗
PubMed1997

Uterine stromal cell suppression of pIgR production by uterine epithelial cells in vitro: a mechanism for regulation of pIgR production.

Previous studies have shown that the polymeric Ig receptor (pIgR) is produced by rat uterine epithelial cells both in vivo and vitro. The expression of the pIgR is regulated by sex hormones and/or cytokines at mucosal sites, however the mechanism of regulation in the uterus is not clear. In these studies, co-culture of stromal cells from mature rat uteri with uterine epithelial cells decreased epithelial cell pIgR production. Conditioned supernatants from stromal cells incubated with epithelial cells also decreased pIgR production. Immunohistochemical studies confirmed that expression of pIgR on uterine epithelial cells decreased in the presence of stromal cells. Viability of epithelial cells was sustained during these experiments, as evidenced by the maintenance of high transepithelial resistance. These studies are the first report of stromal cell regulation of pIgR production by epithelial cells at any site in the body and suggest that stromal cells can provide a signal that leads to the regulation of pIgR production.

Animals↗
PubMed2000

Limitation of nitric oxide production: cells from lymph node and spleen exhibit distinct difference in nitric oxide production.

Many types of cells in the immune system have been found to produce nitric oxide (NO), which performs multiplex functions. However, in myelin basic protein peptide 68-86 (MBP 68-86)-induced experimental autoimmune encephalomyelitis (EAE) in Lewis rats, we found that elevated NO production was generated from spleen cells (SC), not from lymph node cells (LNC). LNC expressed lower NO synthase 2 (NOS2) and produced lower levels of NO than SC upon MBP 68-86 stimulation. Expression of B7-1(CD80) and B7-2(CD86) molecules was much lower on LNC than on SC. Blocking of B7-1 or B7-2 ligation resulted in reduced NO production by SC. Unlike SC, LNC were resistant to interferon-gamma- or lipopolysaccharide-induced NO production. NO derived from SC suppressed cell proliferation and induced apoptosis in vitro. Addition of N(omega)-nitrol-L-arginine methylester (L-NAME) into cell cultures promoted cell expansion and reduced apoptosis. These results indicate that NO production originates from SC, but not from LNC. Low expression of co-stimulatory molecules and NOS2 of LNC limits NO induction. The high levels of NO derived from SC are involved in the self-limiting mechanisms of autoimmune responses by inhibiting cell expansion and promoting cell apoptosis.

Amino Acid Sequence↗
PubMed1999

Involvement of protein kinases in signalling for nitric oxide (NO) and tumour necrosis factor alpha (TNF) production by monocytes stimulated with colorectal DeTa cancer cells: the lack of evidence for the role of TNF in the regulation of NO production.

Kinases that are involved in NO and TNF production by human monocytes (MO) stimulated by colorectal cancer (DeTa) cells and effects of exogenous and endogenously synthesized TNF on NO induction were studied. The results based on the use of various inhibitors of protein kinases suggest that different signalling pathways operate in MO during induction of TNF and NO release after stimulation by DeTa cells. Stimulation of NO production required at least PTK, PKC and PKA, but only PTK and PKC were engaged in signal transduction for TNF production. Exogenous TNF and TNF produced by MO upon contact with DeTa cells was not sufficient for the induction or enhancement of NO synthesis in MO. The TNF synthesis was not influenced by neither exogenous nor endogenous NO produced by MO in the co-culture. Therefore, signal transduction pathways operating in MO during NO induction seem to be different from these engaged in TNF production, and both regulatory pathways probably operate in MO independently.

Carcinoma↗
PubMed2000

The use of a starter culture in the fermentation of cassava for the production of "kivunde", a traditional Tanzanian food product.

Three cassava fermentation methods (spontaneous fermentation, back-slopping and the use of starter culture) for the production of kivunde, executed in three trials at 30 degrees C, were compared in terms of cyanide level reduction, microbiology and product quality improvement. Among the isolates from spontaneously fermented cassava batches, four strains were selected on the basis of their enzymatic activities and acid production. All were identified as Lactobacillus plantarum and were used as starters in this study. Lowest residual cyanide levels were detected after 120 h fermentation time in samples fermented with the starter culture and were below the maximum value of 10 mg/kg recommended by the Codex/FAO for cassava flour. This finding seems to be related to the alpha-glucosidase activity of the inoculated strains of which API-zyme (Bio-Merieux) tests showed activities of between 20 and > or = 40 nmol/4 h. The total residual cyanide levels of the spontaneous and back-slopping fermentations at 96 h were respectively 43.5 and 47.7 mg/kg dry weight of cassava. Extension of the fermentation period to 5 days, lead to further substantial reduction in the residual cyanide level in both these processes, but not below the recommended maximum value as in the case of starter culture fermented products. The spontaneous and back-slopping fermented cassava showed signs of deterioration after 3 days of fermentation. There was a sharp drop of pH and an increase of titratable acidity for all three batches during the first 48 h followed by a slow rise of pH and drop in titratable acidity towards the end of fermentation. The samples fermented with the starter culture had a smooth texture and pleasant fruity aroma, as opposed to the course and dull appearance and more complex flavour of the samples of spontaneous and back-slopping batches. During fermentation with starter culture, Enterobacteriaceae and yeasts and moulds could not be isolated throughout the period of fermentation (detection limit: 10 colony forming units/g). The present findings indicate the suitability of these Lb. plantarum strains as starter cultures for cassava fermentation in the kivunde process. The paper highlights the potential for the improvement of a traditional African fermented food (kivunde) through the use of a starter culture.

Bacteriological Techniques↗
PubMed2003

Metabolic network analysis during fed-batch cultivation of Corynebacterium glutamicum for pantothenic acid production: first quantitative data and analysis of by-product formation.

A first generation genetically modified strain of Corynebacterium glutamicum has been assessed for its potential to synthesise and accumulate the vitamin pantothenic acid in the medium using fed-batch cultivation technology, with biomass concentration controlled by isoleucine limitation. Kinetic analysis of specific rates throughout the process has been used to model carbon flux through both central metabolism and the specific pathways involved in product formation. Flux towards pantothenic acid is potentially high but much of this flux is dissipated as by-products within associated pathways, notably linked to amino acid synthesis. The major limitation of vitamin production in this strain is linked to the tenfold higher flux of keto-isovalerate towards valine rather than pantothenic acid. Attempts to modify this ratio by imposing nitrogen limitation provoked carbon overflow as unidentified non-nitrogenous compounds. The observed accumulation of glycine suggests that the flux towards pantothenate production may by limited by the rate of the pathway intermediate (5,10-methylene-tetrahydrofolate) regeneration.

Bioreactors↗
PubMed1997

Development of an equilibrium immunoassay using electrochemiluminescent detection for a novel recombinant protein product and its application to pre-clinical product development.

In order for a biotechnology derived protein product to be considered 'well characterized', a thorough understanding of the pharmacokinetic and metabolic fate of the product is essential. Specifications for such products need to be based on historical data obtained in the pre-clinical, clinical and manufacturing experience through the use of specific, accurate, precise and validated assays. Typically, assays such as ELISA or RIA that have the disadvantages of limited dynamic range, matrix interactions and hazardous waste generation are used to gather this data. We present data in this abstract that demonstrates the utility of an equilibrium immunoassay that uses electrochemiluminescent detection in the assessment of the pharmacokinetic and metabolic fate of a biotechnology derived product based on either the human (AMG1h) or murine (AMG1m) protein sequence. The assay uses biotinylated antibody in a sandwich format with antibody labeled with the N-hydroxysuccinimide ester of a ruthenium (II) tris-bipyridine chelate for detection in the Origen System. Streptavidin-coated paramagnetic beads are used to capture the antibody-antigen-antibody sandwich complex and facilitate electrochemiluminescent detection. The assay standard curve for AMG1h ranges from a lower limit of quantitation (LOQ) of 2.5 ng ml(-1) to an upper limit of quantitation (ULQ) of 2000 ng ml(-1) with accuracy and precision of not greater then 15% CV and deviation from nominal over the range. The corresponding LOQ (0.5 ng ml(-1)) and ULQ (200 ng ml(-1)) values determined for AMG1m displayed similar accuracy and precision. In addition, we demonstrate that the assay as performed is insensitive to matrix effect up to addition of 7%, of the total reaction volume. General guidelines for developing similar electrochemiluminescent based assays and their applications, advantages and limitations will be discussed.

2,2'-Dipyridyl↗