Assessment of DNA-binding affinity of cholinesterase reactivators and electrophoretic determination of their effect on topoisomerase I and II activity.

In this paper, we describe the biochemical properties and biological activity of a series of cholinesterase reactivators (symmetrical bisquaternary xylene-linked compounds, K106-K114) with ctDNA. The interaction of the studied derivatives with ctDNA was investigated using UV-Vis, fluorescence, CD and LD spectrometry, and electrophoretic and viscometric methods. The binding constants K were estimated to be in the range 1.05 × 10(5)-5.14 × 10(6) M(-1) and the percentage of hypochromism was found to be 10.64-19.28% (from UV-Vis titration). The used methods indicate that the studied samples are groove binders. Electrophoretic methods proved that the studied compounds clearly influence calf thymus Topo I (at 5 µM concentration, except for compounds K107, K111 and K114 which were effective at higher concentrations) and human Topo II (K110 partially inhibited Topo II effects even at 5 µM concentration) activity.

DNA-protective activities of hyperforin and aristoforin.

The aim of this study was to explain the molecular mechanisms of action of hyperforin, a phluoroglucinol derivative found in Hypericum perforatum L. and its more stable derivative aristoforin. DNA-topology assay revealed partial DNA-protective activities of hyperforin and aristoforin against Fe(2+)-induced DNA breaks. In order to assess molecular mechanisms underlying DNA-protective activity, the potential antioxidant activity of hyperforin and aristoforin was investigated using DPPH and OH scavenging assays, reducing power assay and Fe(2+)-chelating assay. We also studied interaction of hyperforin and aristoforin with DNA using established protocols for fluorescence titration. The ability of the studied compounds to relax topoisomerase I with electrophoretic techniques was investigated. The reduction in the fluorescence of hyperforin indicated an interaction between hyperforin and DNA with a binding constant of 0.2×10(8)M(-1). We suggest that a mechanism of hyperforin/aristoforin DNA-protective abilities is based on free radicals (mainly OH) scavenging activity.

Lichen secondary metabolites as DNA-interacting agents.

A series of lichen secondary metabolites (parietin, atranorin, usnic and gyrophoric acid) and their interactions with calf thymus DNA were investigated using molecular biophysics and biochemical methods. The binding constants K were estimated to range from 4.3×10(5) to 2.4×10(7)M(-1) and the percentage of hypochromism was found to be 16-34% (from spectral titration). The results of spectral measurement indicate that the compounds act as effective DNA-interacting agents. Electrophoretic separation studies prove that from all the metabolites tested in this study, only gyrophoric acid exhibited an inhibitory effect on Topo I (25µM).

Study of DNA interactions with cyclic chalcone derivatives by spectroscopic techniques.

A series of chalcone derivatives (1-4) were studied. The interaction between these ligands and calf thymus DNA was studied with UV-vis spectrophotometry, fluorescence and circular dichroism spectroscopy. The binding constants K were estimated at 0.5-4.6×10(5) M(-1). All these measurements indicated that the compounds behave as effective DNA-intercalating agents. Electrophoretic separation proved that ligands inhibited topoisomerase I at a concentration of 60 µM.

Interaction of a copper(II)-Schiff base complexes with calf thymus DNA and their antimicrobial activity.

The interaction of a copper complexes containing Schiff bases with calf thymus (CT) DNA was investigated by spectroscopic methods. UV-vis, fluorescence and CD spectroscopies were conducted to assess their binding ability with CT DNA. The binding constants K have been estimated from 0.8 to 9.1×10(4) M(-1). The percentage of hypochromism is found to be over 70% (from spectral titrations). The results showed that the copper(II) complexes could bind to DNA with an intercalative mode. Synergic action of Cu(II) complexes with ascorbic acid against Candida albicans induced the generation of free radicals and increased (more than 60 times) antimicrobial effect of these complexes.

DNA binding properties and evaluation of cytotoxic activity of 9,10-bis-N-substituted (aminomethyl)anthracenes.

The results of DNA binding properties for four selected N-substituted 9,10-bis(aminomethyl)anthracenes are presented. DNA binding affinities were studied using UV-vis and fluorescence spectrophotometric titrations, CD spectroscopy, denaturation transition temperature (Tm) measurements and AM1 quantum chemical calculations. The results obtained indicate that the anthracene products intercalate into the stacked base pairs of DNA with binding constants, K, in the range 1.3-10.9 x 10(5)M(-1) and the binding site size in DNA-base pairs, n, extending over the range 2.4-4.6. Tm values increased in the presence of the anthryl probes, thereby reflecting an increased stability of the calf-thymus (CT) DNA double helix and rendering agreement with the spectrometric titration results. The synthesized compounds were tested against L1210 and HeLa tumor cell lines wherein the HeLa cells appeared to be more sensitive than the L1210 cells. 9,10-Bis{[2-(piperazin-1-yl)ethyl]aminomethyl}anthracene exhibited the highest activity of the tested compounds. Our findings were compared with those of a control drug bisantrene.

Determination of the binding affinities of plasmid DNA using fluorescent intercalators possessing an acridine skeleton.

Novel acridine derivatives, wherein the steric factor has been varied systematically through substitution at the 9 position of the acridine ring, were evaluated as convenient fluorescent probes for nucleic acid detection. The binding affinities of N-(9-acridinylthiocarbamoyl)amino acids (ATA) with plasmid DNA (pUC 19) were investigated using UV-vis spectrophotometry, fluorometric titration and quantum chemical calculations (AM1). From spectrofluorometric analysis, the binding constants for the DNA-ATA complexes were determined. To elucidate its DNA intercalation, the most preferable tautomeric structure of ATA was established by means of AM1 calculations.

The effect of essentiale on histones and nucleic acids in liver and blood-forming tissues of rats irradiated with gamma-rays.

In this paper, the influence of the hepatoprotective drug Essentiale on the target (normal and regenerating liver) and the non-target (spleen and bone marrow) rat tissues was studied after whole body irradiation with the dose of 5.7 Gy gamma-radiation. The application of the drug 24 h before irradiation alleviated all the radiation induced changes of the histones and nucleic acids in the normal and regenerating liver tissue. In the non-target tissues only mild radioprotective effect was observed. The application of the preparation 30 min after irradiation was less effective than the application before irradiation. The repeated application of Essentiale after irradiation did not increase the beneficial effect of the previous preparation application.

Effect of aging and gamma radiation on acetylation of rat liver histones.

We studied age-related and radiation induced changes in histone acetyltransferase activity and acetylation of histone fractions in normal and regenerating rat livers. Male Wistar rats aged 1-28 months were used through the studies. They were exposed to whole-body doses of 5.7 Gy gamma radiation from a 60Co source and partial hepatectomy was carried out 30 min after irradiation. Within nuclei isolated from normal and regenerating livers of rats, acetyltransferase activity of histones increased with age. An age-dependent decrease in histone acetyltransferase activity in normal and regenerating rat livers was also obvious at the time of maximum activity (i.e. at the 4th min of incubation). The radiation-induced drop in acetyltransferase activity was mild in normal livers. Twenty-four hours after partial hepatectomy, acetyltransferase activity in regenerating livers was almost completely inhibited by irradiation. The acetylation of histone fractions and subfractions was not equal. The highest activity among all histone fractions was found in fraction H4. In regenerating livers of non-irradiated and irradiated rats, there was a greater rate of acetylation in tetraacetylated subfractions of histone H4 compared with intact livers.

[The effect of silymarin, a hepatoprotective substance, on liver histones in irradiated rats]. / Vplyv hepatoprotektívnej látky--silymarínu na históny v peceni oziarených potkanov.

Changes of concentration, total content of histones and relative portions of histone fractions were investigated in the liver of rats after administration of the hepatoprotective substance silymarin (70 mg/kg) and after gamma-irradiation of the whole body at a dose of 3 Gy, which were examined in 30 hours and in 7 days. Administration of silymarin alone considerably increased the concentration, particularly total content of extractable histones in the liver of rats examined in hour 30. They decreased below the level of control values after 7 days. The whole body irradiation at a dose 3 Gy of gamma-radiation caused a steep fall of the concentration and total content of histones in hour 30, which persisted also on day 7. Silymarin administered 1 hour before irradiation prevented quantitative changes of histones in hour 30, after irradiation the fall was still steeper than after irradiation without silymarin administration. As Tab. I shows, a significant decrease in the relative portion of histone fractions H2A+H2B was found in the extracted histone of the experimental animals of all 3 groups in hour 30, as well as a decrease in the fraction H1 after irradiation without silymarin administration. A decrease in the lysin-rich-histone portion was related to an increase in the relative portion of histone H3. In the rats which were administered silymarin 1 hour before irradiation these changes were found to persist until day 7, and they were related to an increase in the subfraction H1(0) within the histone fraction H1 (Tab II). Hence the results document that silymarin administration 1 hour before irradiation had a positive effect which was observed in all the investigated parameters in hour 30 after irradiation. But the radioprotective effect of silymarin was only temporary while until day 7 after irradiation histone variations were identical or still larger than after irradiation without silymarin administration.

Aging and radiation induced alterations in liver histones.

Age-related changes in histones in the liver of normal rats and in rats irradiated with 5.7 Gy gamma rays were examined. Quantitative histone changes in growing and aging rats (from 1 to 28 months of age) were found to be mild only. As they paralleled the DNA changes, the histone/DNA ratio remained stable with age. In total extracted histones there was a decrease in the H1 proportion in older age groups with preceding increase in the H10 proportion. Thirty minutes after irradiation the amount of histones was reduced with age, probably due to an impaired extractability of histones. As the quantitative DNA changes were milder, the histone/DNA ratio decreased in aging liver after irradiation. Similar patterns of changes in proportion of the H1 fraction and H10 subfraction were observed in irradiated and nonirradiated animals, in the former with an earlier onset. Irradiation, therefore, accelerated spontaneous age-related alterations.

Aging and radiation induced alterations of histones in regenerating rat liver.

We studied the age-related and radiation-induced changes in histones in regenerating liver of rats between 1 and 28 months of age. The examinations were carried out 30 h after a two-third hepatectomy, i.e. after the first wave of DNA and histone synthesis. During induced regeneration age-related changes in liver manifested themselves in decreasing the cellularity per gram wet weight, in slowing down the increase in DNA and histone contents per organ, in changing the mutual proportions of histone fractions H1, H2A + H2B, H4 and in increasing the H1 zero variant within histone H1. Radiation-induced latent injury (a dose of 5.7 Gy gamma radiation 30 min before partial hepatectomy) was manifested during induced regeneration in similar but much more profound changes with their earlier onset. Since the changes in regenerating liver were found not be milder than in intact liver it means, that induced proliferation did not lead to elimination of altered cells or to their rejuvenation.

[Changes in liver histones in rats after single, continuous and combined irradiation with gamma rays]. / Zmeny histónov v peceni potkanov po jednorazovom, kontinunálnom a kombinovanom oziarení gama lúcmi.

The authors examined the concentration and total contents of histones and the relative ratio of different fractions of histones in the liver of rats after a single (6 Gy), continuous (total dose 1.4 Gy cumulated at rate of 0.1 Gy per day) and combined continuous and single irradiation with gamma rays. The most extensive changes of histones were revealed after combination of continuous and single irradiation. Continuous irradiation did not lead to activation of reparative mechanisms which would mitigate the response to the subsequent single irradiation. The summation of effects is apparent from the fact that in the liver, contrary to organs with active cellular proliferation, irradiation with small doses does not increase the resistance to the subsequent irradiation, on the contrary, cumulation of damage was observed.