Zusammenhang zwischen den Protein-Thiolen und der Dopplungszeit verschiedener Ratten-Hepatome

Thiol levels were measured in three cell lines derived from rat hepatocytes with different growth rates and degrees of tumorigenicity: IAR20 having normal epithelial morphology and no tumour forming ability; IAR6.1 being a chemically-transformed malignant cell line; and IAR6.1RT7 derived from an epithelial tumour obtained after injection of IAR6.1 cells into a syngenic animal. The mean levels of GSH, GSSG, low molecular weight thiols (LMWT), macromolecular thiols (MT) and total reactive protein sulphur (TRPS), expressed as nmoles-SH mg-1 protein, were found to be 25.5, 7.5, 50.1, 114.5 and 143.6 respectively for IAR20; 37.6, 3.9, 65.4, 126.8 and 148.4 for IAR6.1; 17.2, 4.4, 52.3, 141.0 and 168.2 for IAR6.1RT7. Cultures were treated with D,L-buthionine-S,R-sulphoximine (BSO) to cause greater than 70 per cent depletion of GSH and the measurements of cellular thiols repeated. Although treatment with BSO caused a substantive decrease in the LMWT fraction, there were no major changes in macromolecular thiols or in total reactive protein sulphur. The respective mean values for LMWT, MT and TRPS (expressed as nmoles-SH mg-1 protein) were 19.4, 109.8, 136.3 for IAR20; 17.2, 119.3, 143.6 for IAR6.1; 21.6, 150.7 and 163.5 for IAR6.1RT7. It is concluded that significant differences in thiol levels exist between the three rat liver cell lines studied. However, severe acute depletion of GSH is not reflected by changes in the levels of macromolecular thiols which suggests that there is only a slow equilibrium between these two major thiol pools.

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Cellular thiols in rat liver cell lines possessing different growth characteristics

Principe

Riley

Slater

1991

Cell Biochemistry & Function

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Flow cytometric analysis of protein thiol groups in relation to the cell cycle and the intracellular content of glutathione in rat hepatocytes

Principe¹,

Wilson

Riley³

et al. 1989

Cytometry

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The relationship between protein thiols (PSH) and cell proliferation was examined in ethanol-fixed rat hepatocytes. A new protocol was developed for simultaneous measurement of protein thiol vs. DNA content by flow cytometry. The fluorescent dye o-phthalaldehyde (OPT) was used for flow cytometric measurements of protein thiol groups. The influence of nonprotein thiols was examined by monitoring the cell cycle of cells in which the glutathione content (GSH) was modified by treatment with buthionine sulphoximine (BSO). Three rat liver cell lines (IAR 20, IAR 6.1, IAR 6.1RT7) were used: these cell lines possess different growth characteristics and degrees of tumorigenicity, which made it possible toanalyse changes in PSH during normal and deranged cell proliferation. The effects on the cell cycle of the changes in PSH due to the depletion of GSH were measured by 5-bromo-2'-deoxyuridine (BrdUrd) incorporation and flow cytometry. The data obtained can be summarised as follows: a) OPT fluorescence increases with increasing DNA content in all rat liver cell lines examined; b) the greatest variation in PSH content occurs in GI.There is a smaller variation in Gz + M, and PSH levels are relatively invariant throughout S-phase; c) a higher content of PSH is found in the tumorigenic cell lines; d) the amount and distribution of PSH is not affected by BSO treatment; e) kinetic studies indicate that BSO treatment has no effect on the ability of the IAR rat liver cell lines to progress through the cycle.Key terms: Flow cytometry, reduced glutathione, o-phthalaldehyde (OPT), D,Lbuthionine-S, R-sulphoximine (BSO) Hammett (18,191 proposed that there is a relationship between thiol groups and cell division. Rapkine (36) concluded that there was evidence of a thiol cycle during cell growth and postulated that cell division was associated with a decrease in the oxidation-reduction potential in the cell. Mazia (26) confirmed Rapkine's results and speculated that the interaction between protein and nonprotein thiol levels was important for cell division. These pioneer observations have generated a wide range of investigations on material from different sources, both of a normal and a pathological nature (1,6,12,20,30,39) that have suggested interesting approaches to a better understanding of the mechanism of cell division (4,31,42,44,45).Although these studies have repeatedly suggested that thiol groups may have an important role in cell division, the degree of interaction between the protein and the nonprotein thiol groups remains to be established. Can cell division be triggered by changes in the contents of thiol groups of protein or nonprotein origin? In this paper we present results from a new flow cytometric procedure that has been developed to analyse protein thiol distribution in cells in different phases of the cell cycle by simultaneously measuring cellular protein thiol levels and total DNA content. The study was carried out using three different rat liver cell lines. The parent cell line, IAR 20, was derived from a primar...

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The redox-mediated control of enzyme function and cellular structure

Gascoyne

2009

Int. J. Quantum Chem.

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A theoretical study is presented in which the feasibility of the redox control of enzymic function and cellular structure is demonstrated. It is shown that dithiol-and disulfide-dependent enzymes may be expected to show a strong activity dependence on the oxidation state of the cytoplasmic glutathione in living cells. Cellular structure that is dependent on interprotein disulfide bonds also shows a strong dependence on the glutathione status. Such redox control is only feasible when the cytoplasmic glutathione is at least 99% reduced. A redox control mechanism would require that a cellular machinery exist in order to control the oxidation status of the cytoplasmic glutathione. The existence of such a control machinery is proposed.

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Zusammenhang zwischen den Protein-Thiolen und der Dopplungszeit verschiedener Ratten-Hepatome

Cited by 4 publications

References 13 publications

Cellular thiols in rat liver cell lines possessing different growth characteristics

Cellular thiols in rat liver cell lines possessing different growth characteristics

Flow cytometric analysis of protein thiol groups in relation to the cell cycle and the intracellular content of glutathione in rat hepatocytes

The redox-mediated control of enzyme function and cellular structure

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