Immunospecific protein of 34.5 kDa from DNA-protein cross-links induced by cis- and trans-diamminedichloroplatinum

Cis-diamminedichloroplatinum(II) (cis-DDP) is one of the most often used anticancer drugs. It is generally accepted that the antitumor activity of the drug results from its interactions with DNA. Trans-diamminedichloroplatinum(II) (trans-DDP) also binds to DNA effectively, but is clinically ineffective. In the present work the lymphocyte nuclear proteins that participate in DNA-protein cross-links induced by cis- and trans-DDP are investigated. In lymphocytes which are incubated without platinum compounds there are DNA-binding proteins in the range of 45-71 kDa. It is shown that additional proteins of 28, 30, 34.5, 45 and 120 kDa are cross-linked with DNA in lymphocytes after 2-h incubation with cis-DDP at concentrations of 0.1 and 0.5 mM. Trans-DDP does not bind additional proteins to DNA after the same incubation time. Electrophoretic analysis shows that trans-DDP binds much more of the same nuclear proteins to DNA than cis-DDP after 12-h incubation. In this study a test for the identification of 34.5 kDa protein is also undertaken. This protein appears in the samples obtained after 12-h incubation of lymphocytes with cis- and trans-DDP at 0.5 and 1 mM, especially. The protein of 34.5 kDa from cross-links induced by 1 mM trans-DDP is recognized by antibodies against the protein of the same molecular weight from the nuclear matrix of the lymphocytes. The results obtained here are discussed in relation to the biological activity of diamminedichloroplatinum isomers.     

Morphological and biochemical changes in human fibroblast lines induced by anthracyclines during apoptosis

We show that treating human trisomic fibroblasts with anthracyclines - aclarubicin, daunorubicin and idarubicin - leads to certain changes in these cells; namely the activation of caspase 3, morphological changes and an increase in the level of intracellular calcium. These results suggest that anthracycline drugs are also able to induce apoptosis in pathological, trisomic cells.     

Effects of pyrroline and pyrrolidine nitroxides on lipid peroxidation in heart tissue of rats treated with doxorubicin

Protection from doxorubicin-induced lipid peroxidation in vivo by two pyrroline and pyrrolidine nitroxides, Pirolin, PL, and Pirolid, PD, was examined in the heart tissue of rats treated with this drug. The level of lipid peroxidation was estimated on the basis of MDA content. A considerable (three-fold) increase in the MDA amount was found in heart homogenates from rats injected with doxorubicin, whereas no significant changes in MDA content compared to control were observed in cardiomyocytes treated with the nitroxides (Pirolin or Pirolid) only. Pirolin injected simultaneously with doxorubicin showed antioxidative effect and markedly attenuated lipid peroxidation in the heart tissue caused by this drug. In contrast to Pirolin, structurally related Pirolid was ineffective in the protection of heart myocytes from DOX-induced lipid peroxidation.     

Intraspecies variability of Desulfovibrio desulfuricans strains determined by the genetic profiles

Fifteen (soil and intestinal) strains of Desulfovibrio desulfuricans species were typed by PCR method with the use of primers specific for repetitive extragenic palindromic (REP) and enterobacterial repetitive intergenic consensus (ERIC) sequences. As a result, characteristic DNA fingerprints for the strains were obtained. Moreover, the genetic profiles were found to be useful for typing and distinguishing the strains of D. desulfuricans. According to cluster analysis, PCR with primers complementary to the sequences REP appeared to be slightly more discriminatory than PCR with ERIC primers for the investigated strains. Distinct fingerprint patterns of two isolates derived from the same patient pointed to the different origin of both strains.     

Impaired angiogenesis in neuropeptide Y (NPY)-Y2 receptor knockout mice

Which of Y1-Y5 receptors (Rs) mediate NPY's angiogenic activity was studied using Y2R-null mice and R-specific antagonists. In Y2R-null mice, NPY-induced aortic sprouting and in vivo Matrigel capillary formation were decreased by 50%; Y1R-antagonist blocked the remaining response. NPY-induced sprouting was equally inhibited by Y2R- (and Y5R- but less by Y1R-) antagonists in wild type mice. Spontaneous and NPY-induced revascularization of ischemic gastrocnemius muscles were similarly reduced in Y2R-null mice. Thus, NPY-induced angiogenesis, spontaneous and ischemic, is primarily mediated by Y2Rs. However, Y5Rs and, to a lesser degree Y1Rs, also may play a role in NPY-mediated angiogenesis.     

Testosterone affects hormone-sensitive lipase (HSL) activity and lipid metabolism in the left ventricle

Fatty acids, which are the major cardiac fuel, are derived from lipid droplets stored in cardiomyocytes, among other sources. The heart expresses hormone-sensitive lipase (HSL), which regulates triglycerides (TG) breakdown, and the enzyme is under hormonal control. Evidence obtained from adipose tissue suggests that testosterone regulates HSL activity. To test whether this is also true in the heart, we measured HSL activity in the left ventricle of sedentary male rats that had been treated with testosterone supplementation or orchidectomy with or without testosterone substitution. Left ventricle HSL activity against TG was significantly elevated in intact rats supplemented with testosterone. HSL activity against both TG and diacylglyceride was reduced by orchidectomy, whereas testosterone replacement fully reversed this effect. Moreover, testosterone increased left ventricle free fatty acid levels, caused an inhibitory effect on carbohydrate metabolism in the heart, and elevated left ventricular phosphocreatine and ATP levels as compared to control rats. These data indicate that testosterone is involved in cardiac HSL activity regulation which, in turn, may affect cardiac lipid and carbohydrate metabolism.     

Effect of mitochondrial genome rearrangement on respiratory activity, photosynthesis, photorespiration and energy status of MSC16 cucumber (Cucumis sativus) mutant

The effects of changes in mitochondrial DNA in cucumber ( Cucumis sativus L.) mosaic mutant (MSC16) on respiration, photosynthesis and photorespiration were analyzed under non-stressed conditions. Decreased respiratory capacity of complex I in MSC16 mitochondria was indicated by lower respiration rates of intact mitochondria with malate and by rotenone-inhibited NADH or malate oxidation in the presence of alamethicin. Moreover, blue native PAGE indicated decreased intensity of protein bands of respiratory chain complex I in MSC16 leaves. Concerning the redox state, complex I impairment could be compensated to some extent by increased external NADH dehydrogenases (ND_exNADH) and alternative oxidase (AOX) capacity, the latter presenting differential expression in the light and in the dark. Although MSC16 mitochondria have a higher AOX protein level and an increased capacity, the AOX activity measured in the dark conditions by oxygen discrimination technique is similar to that in wild-type (WT) plants. Photosynthesis induction by light followed different patterns in WT and MSC16, suggesting changes in feedback chloroplast ΔpH caused by different adenylate levels. At steady-state, net photosynthesis was only slightly impaired in MSC16 mutants, while photorespiration rate (PR) was significantly increased. This was the result of large decreases in both stomatal and mesophyll conductance to CO₂, which resulted in a lower CO₂ concentration in the chloroplasts. The observed changes on CO₂ diffusion caused by mitochondrial mutations open a whole new view of interaction between organelle metabolism and whole tissue physiology. The sum of all the described changes in photosynthetic and respiratory metabolism resulted in a lower ATP availability and a slower plant growth.