Culture experimental system

B Non-human effects experimental observations In toxicologic testing In animals or In bacterial or cell culture test systems ... 

Precellular solute ionization dictates membrane permeability dependence on mucosal pH. Therefore, lumenal or cellular events that affect mucosal microclimate pH may alter the membrane transport of ionizable solutes. The mucosal microclimate pH is defined by a region in the neighborhood of the mucosal membrane in which pH is lower than in the lumenal fluid. This is the result of proton secretion by the enterocytes, for which outward diffusion is slowed by intestinal mucus. (In fact, mucosal secretion of any ion coupled with mucus-restricted diffusion will provide an ionic microclimate.) Important differences in solute transport between experimental systems may be due to differences in intestinal ions and mucus secretion. It might be anticipated that microclimate pH effects would be less pronounced in epithelial cell culture (devoid of goblet cells) transport studies than in whole intestinal tissue. 

Attempts to study the entry of ES products into cells using markers of fluid phase endocytosis yielded unexpected results. When larvae browse resistant IEC-6 cells in the presence of extracellular fluorescent dextran, dextran enters the cytoplasm of a significant proportion of the cells in the mono-layer (Butcher et al., 2000). The parameters of dextran entry are most compatible with the conclusion that larvae wound the plasma membranes of IEC-6 cells that is, they create transient breaches in the membrane that allow impermeant markers to enter the cell (McNeil and Ito, 1989). Wounding is considered to be a common occurrence in intestinal epithelia (McNeil and Ito, 1989). Injured cells are able to heal their wounds by recruiting vesicles to seal the breach (Steinhardt et al., 1994). In an experimental system, healing allows the injured cell to retain cytoplasmic dextran. In epithelial cell cultures inoculated with T. spiralis larvae, the relationship between glycoprotein delivery and injury of plasma membranes is not clear, i.e. dextran-laden cells do not always stain with Tyv-specific antibodies and... 

Both active and passive transport occur simultaneously, and their quantitative roles differ at different concentration gradients. At low substrate concentrations, active transport plays a major role, whilst above the concentration of saturation passive diffusion is the major transport process. This very simple rule can be studied in an experimental system using cell culture-based models, and the concentration dependency of the transport of a compound as well as asymmetric transport over the membrane are two factors used to evaluate the presence and influence of transporters. Previous data have indicated that the permeability of actively absorbed compounds may be underestimated in the Caco-2 model due to a lack of (or low) expression of some uptake transporters. However, many data which show a lack of influence of transporters are usually derived from experiments... 

Biological. Under aerobic conditions or in experimental systems containing mixed cultures, hexachloroethane was reported to degrade to tetrachloroethane (Vogel et al, 1987). In an uninhibited anoxic-sediment water suspension, hexachloroethane degraded to tetrachloroethylene. The reported half-life for this transformation was 19.7 min (Jafvert and Wolfe, 1987). When hexachloroethane (5 and 10 mg/L) was statically incubated in the dark at 25 °C with yeast extract and settled domestic wastewater inoculum for 7 d, 100% biodegradation with rapid adaptation was observed (Tabak et al, 1981). 

Figure 19.7 Experimental procedure forthe assessment of photocytotoxicity (MTT assay) and photo-genotoxicity (comet assay) on reconstructed epidermis. Drugsorformulationscan be applied on the skin surface (topical route) or provided in the culture medium (systemic route see [76]).

Hence, these Qc values are a quantitative measure for the relative affinities of the various NACs to the reactive sites. Figs. 14.10e and/show plots of log Qc versus h(AtN02)/0.059 V of the 10 monosubstituted benzenes. A virtually identical picture was obtained for the log Qc values derived from an aquifer solid column and from a column containing FeOOH-coated sand and a culture of the iron-reducing bacterium, Geobacter metallireducens (GS15). Furthermore, a similar pattern (Fig. 14.10c) was found when correlating relative initial pseudo-first-order rate constants determined for NAC reduction by Fe(II) species adsorbed to iron oxide surfaces (Fig. 14.12) or pseudo-first-order reaction constants for reaction with an iron porphyrin (data not shown see Schwarzenbach et al., 1990). Fig. 14.12 shows that Fe(II) species adsorbed to iron oxide surfaces are very potent reductants, at least for NACs tv2 of a few minutes in the experimental system considered). 

Hjortso, M. A. and H. E. Flores, "Root Culture as an Experimental System for the Production of Plant Chemicals," Paper presented at AIChE 1987 Annual Meeting, New York, NY, November 15-20,1987. 

A schematic of the experimental system is shown in Fig. 1. One liter of culture liquid was added to a 2.8-L cylindrical glass column reactor, and 300 mL of cylindrical carrier was packed with rock wool. All of the tubing connections, stoppers, and seals in the column were made of butyl rubber. To prevent photodecomposition of vitamin B12, the outside of the whole device was covered with a vinyl sheet to shut out light. The digestion was carried out at 55°C and 20 d of hydraulic retention time (HRT). 

Second There is no way to detect and measure the rate of occurrence of human germinal mutations. We must rely on information from experimental organisms and cell-culture test systems and on the belief that these tests mimic what happens in human germ cells. 

As trial system to test the application of the proposed model the ability of encapsulated XAD-7 was evaluated for the selective separation of berberine from dilute aqueous mixtures of berberine and dopamine, the target secondary metabolite, and an undesirable intermediate metabolite of Thalictrum rugosum plant cell culture [18]. Competitive adsorption experiments were performed in dilute aqueous mixtures of berberine and dopamine, both at initial concentrations of 60 mg l-1, which is representative of actual plant cell culture. Experimental and theoretical results for normalized bulk concentration profiles of berberine and dopamine are shown in Fig. 10. The bulk berberine concentration was reduced to approximately 4.6% of the initial concentration, which indicates that 95.4% of the berberine in the initial mixed solution was adsorbed. Encapsulated XAD-7, therefore, selectively concentrated the berberine from dilute aqueous mixtures of berberine and dopamine. 

Immortalized hepatocytes A major drawback of the use of primary hepatocytes is that hepatocytes can not be expanded in culture. To overcome this problem, researchers have embarked on the immortalization of primary hepatocyte cultures (e.g. Bayad et al. 1991 Li et al. 2005). Currently, immortalized human hepatocyte cell lines are available commercially and may represent convenient screening experimental systems for enzyme induction studies. Unfortunately, at the time of this writing, there are no peer-reviewed publications on the application of human immortalized hepatocytes in induction studies. It is important to ensure that the induced isoforms in the cell lines are the mature P450 isoforms rather than the embryonic forms. For instance, the use of HepG2 cells may not be appropriate as the embryonic P450 isoforms CYP1A1... 

Cultures of established cell lines or cell strains should be used. Mammalian cell lines should be determined to be mycoplasm-free and should be karyotyped. This is to ensure that they will respond in the expected fashion in the experimental system being used. 

Several experimental systems to check the inhibition potency of bile add transport have been characterized. Using sandwich-cultured human hepatocytes, bosentan, cyclosporin A, CI-1034 (endothelin-A receptor antagonist), glyburide, erythromycin estolate, and troleandomycin could inhibit the taurocholate efflux to the bile pocket [234]. Moreover, Mita et al. [235] construded NTCP/BSEP double-transfeded cells and some cholestasis-induced compounds inhibited both the NTCP-mediated uptake and the BSEP-mediated efflux of taurocholate. Then, they have found fluorescent bile acids whose transcellular transport was dearly observed, which may be used for the rapid identification of inhibitors of NTCP and BSEP in drug screening process [235]. 

Commonly, restrictions are placed on the transport fluxes (corresponding to the various uptake processes). For example, when specific information regarding an uptake rate is not known, the maximal uptake rate is set by using the constraints defined by Eq. (10) (fi is defined as the maximal uptake rate or b. element). However, often in experimental systems, the uptake rates have been measured, and the flux value can be fixed by using the formalism described by Eq. (10). Transport reactions for metabolites that are not present in a simulated culture condition are constrained to zero (0 < vj < 0). 

By definition, the experimental unit is the smallest unit randomly allocated to a distinct level of a treatment factor. Note that if there is no randomization, there is no experimental unit and (in nearly all cases) no experiment. Although it is possible to perform experiments without randomization, it is difficult to do well, and risky unless the experimental system is very well understood (7). Randomization is important for several reasons. Randomization changes the sources of bias into sources of variation in general, a noisy assay is better than a biased assay. Further, randomization allows estimates of variation to represent variation in the population this in turn justifies statistical inference (standard errors, confidence intervals, etc.). A common practice in cell-culture bioassay is to rotate among a small collection of layouts rather than use random allocation. Whereas rotation among a collection of layouts is certainly better than a fixed layout, it is both possible and practical to use carefully structured randomization on a routine basis, particularly when using a robot. 

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