Cytofluorometric determination of protein‐bound thiols and DNA in cell nuclei

Abstract

The aim of the present study was to establish a cytofluorometric method for the simultaneous determination of protein-bound sulfhydryl-groups (PSH) and DNA in isolated cell nuclei. DNA was stained with ethidiumbromide and PSH with N-iodoacetyl-N(5-sulfo-1-naphthyl) ethylendiamine (AEDANS). Disulfide groups of nuclear proteins were determined by the same method after reduction with sodium borohydride or thioglycollic acid. The method was established by using nuclei of human lymphocytes, which then served as a biological standard for further investigations of the nuclei of different mammalian cell types: nuclei from mouse liver cells and nuclei from the cells of two human melanoma cell lines. For non-proliferating lymphocytes distinct DNA- and PSH-values could be measured. The PSH-values detected in the nuclei of the other cell types were higher by comparison and varied within the cell cycle; i.e., PSH increased during the S-phase and was almost doubled during the cell generation cycle from G1- to G2-phase. Cell line and cell cycle-dependent variations of nuclear disulfides could also be detected. These results are discussed with respect to their radiobiological implications. In conclusion, thiol groups may represent one factor determining the radiosensitivity of cells, but they are not the only decisive one.

Sodium thioglycollate enhances pollen germination and pollen tube elongation in cruciferous species

“…Based on the long history of incorporating reducing agents such as mercaptoethanol, dithiothreitol, and thioglycolic acid in extraction buffers to minimize or delay damaging effects of oxidative enzymes on membranes, organelles, proteins, or viruses of interest (Anderson and Rowan 1967;Davis and Hampton 1986;Lau et al 1981;Schabronath et al 1990), our results suggest a protective effect of sodium thioglycollate on pollen physiological functions that enhance both pollen germination and tube elongation. Conceivably, this may occur via different mechanisms, e.g., by initially fostering phospholipid membrane integrity and minimizing leakiness during germination of the pollen on the stigma and, secondly, by creating a less aerobic environment within and around pollen tubes during their elongation in the style.…”

“…Based on our assumption that oxygen tension within the tissue of the style would be lower than in ambient conditions on the stigma, we hypothesized that providing a "reducing" culture environment by addition of the reducing agent sodium thioglycollate (C 2 H 3 O 2 SNa) to the culture medium would enhance pollen tube elongation. Sodium thioglycollate is commonly used in microbiological media to create less aerobic conditions, as well as in plant, microbial, animal, and human cell extraction buffers, to inhibit oxidative reactions (damaging to phospholipids, organelles, cells, or conformations of proteins) and also to optimize plant and fungal virus yields (Anderson and Rowan 1967;Wildes et al 1969;Lau et al 1981;Davis and Hampton 1986;Schabronath et al 1990;Zhong et al 2004). However, to our knowledge, sodium thioglycollate has not been reported to be used in pollen germination media.…”

Programmed self-assembly of enzyme activity-inhibited nanomedicine for augmenting chemodynamic tumor nanotherapy

“…Among them, some genes involved in the DNA replication maintenance of fidelity ( e.g. , PCNA, EXD2, DDX11 and CENPX) and meiosis II cell cycle process 63 ( e.g. , HORMAD2, RAD21, RAD51C and SGO1) were significantly down-regulated after TGA–Cu NPs treatment.…”

“…S23 †). Among them, some genes involved in the DNA replication maintenance of fidelity (e.g., PCNA, EXD2, DDX11 and CENPX) and meiosis II cell cycle process 63 (e.g., HORMAD2, RAD21, RAD51C and SGO1) were significantly down-regulated after TGA-Cu NPs treatment. These genes significantly inhibit the rapid proliferation of tumor cells by affecting the activity of proteins in the centromere region of the chromosome.…”