The avian cell line AGE1.CR.pIX characterized by metabolic flux analysis

Lohr V, Haedicke O, Genzel Y, et al. The avian cell line AGE1.CR.pIX characterized by metabolic flux analysis. BMC Biotechnology. 2014;14(1): 72.Background: In human vaccine manufacturing some pathogens such as Modified Vaccinia Virus Ankara, measles, mumps virus as well as influenza viruses are still produced on primary material derived from embryonated chicken eggs. Processes depending on primary cell culture, however, are difficult to adapt to modern vaccine production. Therefore, we derived previously a continuous suspension cell line, AGE1.CR.pIX, from muscovy duck and established chemically-defined media for virus propagation. Results: To better understand vaccine production processes, we developed a stoichiometric model of the central metabolism of AGE1.CR.pIX cells and applied flux variability and metabolic flux analysis. Results were compared to literature dealing with mammalian and insect cell culture metabolism focusing on the question whether cultured avian cells differ in metabolism. Qualitatively, the observed flux distribution of this avian cell line was similar to distributions found for mammalian cell lines (e.g. CHO, MDCK cells). In particular, glucose was catabolized inefficiently and glycolysis and TCA cycle seem to be only weakly connected. Conclusions: A distinguishing feature of the avian cell line is that glutaminolysis plays only a minor role in energy generation and production of precursors, resulting in low extracellular ammonia concentrations. This metabolic flux study is the first for a continuous avian cell line. It provides a basis for further metabolic analyses to exploit the biotechnological potential of avian and vertebrate cell lines and to develop specific optimized cell culture processes, e.g. vaccine production processes

Plasma membrane receptor for beta-lactoglobulin and retinol-binding protein in murine hybridomas

The aim of the present work was to study the binding of [I-125]-BLGA (beta-lactoglobulin variant A) to the plasma membrane fraction of hybrid cells. This binding increased as a function of time with on-rate and off-rate constant at 4.47 +/- 0.18 x 10(6) M-1 min(-1) and 0.17 +/- 0.07 min(-1), respectively (n = 3). The saturation study showed a single binding site type corresponding to a K-d at 8.26 +/- 2.98 nM and 14.02 +/- 2.61 x 10(12) sites per mg of the plasma membrane protein (n = 3). Competitive of binding BLGA was observed with BLGA, complexed with retinol and also with RBP (retinol-binding protein). Gel filtration of [I-125]-BLGA incubated with Triton X-100 solubilized membrane showed the formation of a ligand-receptor complex. Cross-linking of the tracer to plasma membrane showed a complex with a M-r at 69 kDa, suggesting a receptor M-r of 51 kDa, as seen by autoradiography of SDS-PAGE

Lactosérum et beta-lactoglobuline: 2 coproduits du lait qui peuvent remplacer le sérum de veau foetal dans la culture de cellules d'hybridomes de souris

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PENGARUH EKSTRAK DIKLOROMETAN JAHE (Zinger officinale Roscoe) TERHADAP PENGIKATAN TOKSIN KOLERA B-SUBUNIT CONJUGASI (FITC) PADA RESEPTOR SEL HIBRIDOMA LV DAN CACO-2

Cholera toxin is main factor that responsibility of watery diarrhea. The objectives of this investigation was to study the effect of ginger extract (GDE) in blocking cholera toxin binding to receptors of LV hybridome and Caco-2 cells. Analysis of toxin binding by flow cytometry of 0-5 Æ’ÃÂg/ml of B-FITC conjugated cholera toxin to 105 cells/ml of LV hybridome and Caco-2 cells in RPMI, at 4 ¾aC for one hour showed that specific interaction of B-FITC by 44,44 ¾b 3,49 percent to LV hybridome and by 94,58 ¾b 1,83 percent to Caco-2 cells. A total of 105 cells/ml of hybridome and Caco-2 cells were incubated with 0-5 Æ’ÃÂg/ml toxin B-FITC and 25 or 50 Æ’ÃÂg/ml GDE in RPMI, at 4 ¾aC for one hour showed that the addition of GDE inhibited the toxin binding. The binding inhibition respectively were 4.76-15.66 and 12.55-24.60 percent to Caco-2 cells, and 3.55-17.95 and 3.58-27.83 percent to hybridome cells. The inhibition on the toxin binding may be due to modification of the receptor by GDE or interaction of GDE with the toxin.Key words :  Hibridoma LV, Caco2, ekstrak diklorometan jahe,  and  toksin kolera B-FIT

PENGARUH EKSTRAK DIKLOROMETAN JAHE (Zinger officinale Roscoe) TERHADAP PENGIKATAN TOKSIN KOLERA B-SUBUNIT CONJUGASI (FITC) PADA RESEPTOR SEL HIBRIDOMA LV DAN CACO-2

Cholera toxin is main factor that responsibility of watery diarrhea. The objectives of this investigation was to study the effect of ginger extract (GDE) in blocking cholera toxin binding to receptors of LV hybridome and Caco-2 cells. Analysis of toxin binding by flow cytometry of 0-5 Æ’ÃÂg/ml of B-FITC conjugated cholera toxin to 105 cells/ml of LV hybridome and Caco-2 cells in RPMI, at 4 ¾aC for one hour showed that specific interaction of B-FITC by 44,44 ¾b 3,49 percent to LV hybridome and by 94,58 ¾b 1,83 percent to Caco-2 cells. A total of 105 cells/ml of hybridome and Caco-2 cells were incubated with 0-5 Æ’ÃÂg/ml toxin B-FITC and 25 or 50 Æ’ÃÂg/ml GDE in RPMI, at 4 ¾aC for one hour showed that the addition of GDE inhibited the toxin binding. The binding inhibition respectively were 4.76-15.66 and 12.55-24.60 percent to Caco-2 cells, and 3.55-17.95 and 3.58-27.83 percent to hybridome cells. The inhibition on the toxin binding may be due to modification of the receptor by GDE or interaction of GDE with the toxin.Key words :  Hibridoma LV, Caco2, ekstrak diklorometan jahe,  and  toksin kolera B-FIT

Extracellular Matrix Turnover and Inflammatory Markers Independently Predict Functional Status and Outcome in Chronic Heart Failure

Background Inflammatory pathways may promote extracellular matrix (ECM) remodeling and chronic heart failure (CHF) progression. The relationship between markers of inflammation and of ECM remodeling, and their influence on functional status and outcomes has not been examined in a large cohort of CHF patients. Methods and Results We measured baseline blood serum collagen (amino-terminal propeptide of collagen III [PIIINP], metalloproteinase 1 [MMP-1], tissue inhibitor of metalloproteinase 1 [TIMP-1]), and inflammatory (high-sensitivity C-reactive protein [(hsCRP], interleukin [IL]-18, IL-10) markers in 1009 patients enrolled in the Research into Etanercept Cytokine Antagonism in Ventricular Dysfunction (RECOVER) trial. A positive correlation was detected between the 2 classes of markers (PIIINP to IL-18, MMP-1 and TIMP-1 to CRP, TIMP-1 to IL-18, MMP-1 to IL-10). In the adjusted multivariable model including all biomarkers, only PIIINP (P = .03) and MMP-1 (P = .048) were independent predictors of 6-minute walk test (6-MWT), whereas in another model including only inflammatory biomarkers, IL-18 was an independent predictor. PIIINP (P = .001) was the only biomarker independently associated with death and CHF hospitalization. Conclusions The independent associations of PIIINP and MMP-1 with 6-MWT and PIIINP with CHF morbi-mortality suggest that excessive ECM turnover may be associated with functional capacity deterioration and poor outcome