Assessing cell damage

There are several assays available to quantitate cell death in bioreactors. The simplest assay uses Trypan blue dye exclusion from intact cells to determine hve and dead cell concentrations. Cells that have died and lysed, however, cannot be accounted for using this assay (see Chapter 2, section 2.2). 

Cell damage under different agitation conditions can be assessed by monitoring the increase in LDH release. To equate the LDH content in culture supernatant to the number of cells that have lysed, a standard curve must be established for each cell type and (where applicable) for different environmental conditions to which the cells were exposed. The procedures for the Trypan blue dye exclusion assay and the LDH assay are described in Chapter 2, sections 2.2 and 2.5. 

Cell and Tissue Culture Laboratory Procedures in Biotechnology, edited by A. Doyle and J.B. Griffiths. 1998 John Wiley Sons Ltd. 

Literature reports have used the following reactor parameters to correlate the effects of agitation intensity with cell injury in bioreactors agitator rpm, impeller tip speed, integrated shear factor and Kolmogorov eddy size. Additional parameters have been used for microcarrier bioreactors (discussed below). All correlations of cell injury with a bioreactor parameter should be used only qualitatively. These correlations are, at present, indicative of various trends or mechanistic hypotheses and should not be used for quantitative bioreactor scale-up. In addition, such correlations are applicable to the specific cell type, because different cell types are likely to exhibit different responses to fluid forces. 

The integrated shear factor (ISF), which is assumed to be (incorrectly, strictly speaking) a measure of the strength of the shear field between the impeller and the vessel wall, and may be somewhat more useful for scale-up purposes, is defined as  

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The effects of dmgs and adjuvants must be assessed, both in short-term administration and during chronic treatment. Local effects include changes in mucocihary clearance, cell damage, and irritation. Chronic erosion of the mucous membrane may lead to inflammation, hyperplasia, metaplasia, and deterioration of normal nasal function (76). 

For pharmacological, toxicological and transport studies it is of utmost importance to assess not only the viability but also the functionality of the liver slices. This is essential both for end-point determination of toxic cell damage, and to assess the quality of the tissue during in-... 

In the vestibnlar organ, the initial hair cell damage occurs in the apex of the cristae and the striolar regions of the maculi. Hair cell loss then progresses toward the periphery of the vestibular receptor organ with type I hair cells affected earlier than the type 11 hair cells. Vestibular disturbances from systemic aminoglycoside administration present as severe unsteadiness that becomes worse in the dark, bnt an objective clinical assessment is difficult. In contrast to the situation in the cochlea, regeneration of vestibular hair cells has been observed in mammalian species. ... 

One of the two major tasks of the Committee is to determine how data from diverse test systems can best be used to assess mutagenic damage to human germ cells. 

Fernandez, P.P., Prestamo, G., Otero, L., and Sanz, P.D. 2006. Assessment of cell damage in high pressure shift frozen broccoli Comparison with market samples. Zeitschrift Fiir Lebensmittel-Untersuchung und - Forschung A 224 101-107. 

The flavonol quercetin has been the subject of much interest in terms of its beneficial properties against oxidative stress [62] and has been shown to exert a potent protective actions against hydrogen peroxide-induced apoptosis of rat thymocytes [63] and cell death in rat hepatocytes (BL-9) [60], as well as reducing oxidative stress and cell damage in fiver tumor cells induced by AAPH (2,2-azobis(2-aminopropane) dihydrochloride) [64]. Quercetin and another flavonol, kaempferol, as well as catechin and the flavone taxifolin, have been observed to suppress the cytotoxicity of 02 and H2O2 to Chinese hamster V79 cells, as assessed by the ability of the flavonoids to prevent the decrease in the number of cell colonies induced by the oxidants [65]. Furthermore, quercetin has been shown to protect cutaneous tissue-associated cells (human skin fibroblasts, keratinocytes, and endothelial cells) from oxidative injury induced by buthionine sulfoximine (BSO), an inhibitor of GSH synthesis [66]. 

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