Abstract
Estramustine, a combination of 17 beta-oestradiol and nor-nitrogen mustard, has been shown to be metabolised and to induce specific antiproliferative effects in malignant glioma, including arrest of glioma cells in the G2/M phase of the cell cycle, damage to cell membranes and DNA and induction of free oxygen radicals. To evaluate further the effects of estramustine, an in vivo rat glioma model using inbred BD-IX rats and the BT4C cell line was set up. In order to detect cells with fragmented DNA, tumour and brain specimens were, following fixation for histological examination, processed for in situ end labelling (ISEL) with biotin-labelled nucleotides. Fresh tissue fragments were also used for DNA integrity analysis on agarose gels. It was demonstrated that estramustine induced clusters of ISEL-positive cells and a pronounced typical fragmentation of DNA 0.5-8 h after treatment. In tumours examined 24 or 94 h after estramustine treatment, and in untreated tumours, only occasional single ISEL-positive cells were scattered in the tumour. DNA from normal brain tissue did not display any visible sign of fragmentation. These changes are indicative of programmed cell death induced by estramustine in glioma cells but not in normal brain tissue. Further studies are, however, needed to establish in detail the mechanism of cell death following treatment with the antimitotic drug estramustine.
Highlights
The malignant glioma cells were grown as a monolayer for 1 week before implantation
The digestion was stopped by washing tumours occasional single in situ end labelling (ISEL)-positive cells with morphoseveral times in tap water and washed in buffer for logically fragmented nuclei were seen scattered in the tumour. 5 min
The sections were incubated for 1 h at At 0.5, 1, 2, 4, or 8 h after estramustine treatment clusters of 15°C with buffer containing 0.01 mM dATP, dGTP, dCTP ISEL-positive cells were observed in the tumours (Figure 1). and 0.01 mM biotin dUTP (Boehringer Mannheim, Mann- The tumour tissue between these clusters contained only a heim, Germany) along with 4 U ml-' DNA polymerase 1 few labelled cells
Summary
A rat glioma model using inbred BD- IX rats and the BT4C glioma cell line was set up (Bergenheim et al, 1994). The malignant glioma cells were grown as a monolayer for 1 week before implantation. BD IX rats, 8-14 weeks old, were anaesthetised by i.p. administration of 1.8 ml kg-' of a 1:1 mixture of Hypnorm A microsyringe was used (22S Ga needle; Unimetrics, Shorewood, IL, USA), allowing at least 5 min for injection and withdrawal of the needle to prevent cellular reflux and extracerebral spread of tumour cells. To ensure cell viability the cell suspension was kept on ice during the implantation procedure and the viability of the cell suspension was continuously controlled by staining with trypan blue. After implantation the rats were fed ad libitum
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
