Inhibition of DNA synthesis in human gliomas by roscovitine.

The early effect of 1-100 microM roscovitine, a purine analogue and cyclin-dependent kinase inhibitor, was studied on tissue specimens from eight human malignant gliomas. The tissue was incubated immediately after resection with DMEM containing [3H]methylthymidine plus vehicle alone or the proper concentration of roscovitine for 30-90 min. The DNA synthesis rate was assessed by measurement of [3H]methylthymidine incorporation into trichloroacetic acid insoluble material/mg protein/min. In all gliomas, 100 microM roscovitine inhibited DNA synthesis by 71-97% (average 89 +/- 8%, p<0.0001). This inhibitory effect of roscovitine appeared within 30 min of incubation and was concentration dependent.

Early effects of protein kinase modulators on DNA synthesis in rat cerebral cortex.

By using tissue miniunits, protein kinase modulators, and topoisomerase inhibitors in short-term incubation (0-90 min) we studied (1) the role of protein phosphorylation in the immediate control of DNA replication in the developing rat cerebral cortex and (2) the mechanism of action for genistein-mediated DNA synthesis inhibition. Genistein decreased the DNA synthesis within less than 30 min. None of the other protein kinase inhibitors examined (herbimycin A, staurosporine, calphostin-C) or the protein phosphatase inhibitor sodium orthovanadate inhibited DNA synthesis and they did not affect the genistein-mediated inhibition. The selective topoisomerase inhibitors camptothecin and etoposide decreased the DNA synthesis to an extent similar to that of genistein and within less than 30 min. In addition, the effects of these substances on topoisomerase I and II were studied. Etoposide and genistein but not herbimycin A, staurosporine, or calphostin-C strongly inhibited the activity of topoisomerase II. Our results (1) strongly suggest that the net rate of DNA replication during the S phase of the cell cycle is independent of protein phosphorylation and (2) indicate that the early inhibitory effect of genistein on DNA synthesis is mediated by topoisomerase II inhibition rather than protein tyrosine kinase inhibition.

Development modulates the serum induced effect on the incorporation of [2-3H]mannose into chick optic lobe protein: the possible role of glia.

Recently, we described that serum decreases tritiated mannose incorporation into protein in the chick optic lobe at 18 days of embryonic age (Rossi et al., 1990). In this paper, we found a strikingly different response of this serum effect according to age. The data obtained showed no serum induced decrease in 6-10-day-old embryo. In addition, our results demonstrate that the differential response of the tissue to the serum is independent of the rate of sugar entry into nerve cells. Furthermore, we also report that the variation of mannose or leucine incorporation into protein coincides very closely with the pattern of protein and glycoprotein accumulation during chick optic lobe development. Finally, data were obtained to define glial cells as the cellular target of the serum induced effect. This finding may contribute to elucidate the mechanism of cellular pathogenesis of cerebral lesions that occur after the breakdown of the blood brain barrier, such as in some diseases or during bleeding after injuries.