RESUMO
Here, we use single molecule force spectroscopy performed with optical tweezers in order to investigate the interaction between Caffeine and the DNA molecule for various different concentrations of the alkaloid and under two distinct ionic strengths of the surrounding buffer. We were able to determine the mechanical changes induced on the double-helix structure due to Caffeine binding, the binding mode and the binding parameters of the interaction. The results obtained show that Caffeine binds to DNA by outside the double-helix with a higher affinity at lower ionic strengths. On the other hand, a considerable cooperativity was found only for sufficient high ionic strengths, suggesting that Caffeine may binding forming dimers and/or trimers along the double-helix under this condition. Finally, it was also shown that Caffeine stabilizes the DNA double-helix upon binding, preventing force-induced DNA melting.
Assuntos
Cafeína/farmacologia , DNA/química , Conformação de Ácido Nucleico/efeitos dos fármacos , Algoritmos , Cafeína/química , Modelos Teóricos , Estrutura Molecular , Desnaturação de Ácido Nucleico/efeitos dos fármacos , Pinças Ópticas , Análise EspectralRESUMO
Toxoplasma gondii, the etiological agent of toxoplasmosis, infects nucleated cells and then resides and multiplies within a parasitophorous vacuole. For this purpose, the parasite secretes many virulence factors for the purpose of invading and subverting the host microbicidal defenses in order to facilitate its survival in the intracellular milieu. Essential metals are structural components of proteins and enzymes or cofactors of enzymatic reactions responsible for these parasitic survival mechanisms. However, an excess of non-essential or essential metals can lead to parasite death. Thus, infected host cells were incubated with 20 µM ZnCl2 in conjunction with 3 µM CdCl2 or HgCl2 for 12 h in order to investigate cellular events and organelle damage related to intracellular parasite death and elimination. In the presence of these metals, the tachyzoites undergo lipid uptake and transport impairment, functional and structural mitochondrial disorders, DNA condensation, and acidification of the parasitophorous vacuole, thus leading to parasite death. Additional research has suggested that lysosome-vacuole fusion was involved in parasite elimination since acid phosphatases were found inside the parasitophorous vacuole, and vacuoles containing parasites were also positive for autophagy. In conclusion, low concentrations of CdCl2, HgCl2, and ZnCl2 can cause damage to Toxoplasma gondii organelles, leading to loss of viability, organelle death, and elimination without causing toxic effects to host cells.
Assuntos
Cloreto de Cádmio/farmacologia , Cloretos/farmacologia , Cloreto de Mercúrio/farmacologia , Toxoplasma/efeitos dos fármacos , Vacúolos/efeitos dos fármacos , Compostos de Zinco/farmacologia , Animais , Autofagia , Transporte Biológico/efeitos dos fármacos , Linhagem Celular , Interações Hospedeiro-Parasita/efeitos dos fármacos , Lisossomos , Macaca mulatta , Mitocôndrias/patologia , Desnaturação de Ácido Nucleico/efeitos dos fármacos , Toxoplasma/patogenicidade , Vacúolos/parasitologia , Vacúolos/patologia , Fatores de VirulênciaRESUMO
In Mexico, as in many other countries, traditional medicine is used for the treatment of several diseases. In particular, Amphipterygium adstringens infusion is used for gastritis, gastric ulcers, and gastric cancer. Extracts from this tree have microbicidal effects against Helicobacter pylori, an important risk factor for gastric cancer development. Anacardic acids are constituents of A. adstringens, and 6-pentadecyl salicylic acid (6-PSA) is the most abundant. However, there is a lack of information regarding the effects of 6-PSA on cancer cells. Therefore, we investigated whether 6-PSA has differential effects on the induction of genotoxicity, cytostaticity, and apoptosis in normal human peripheral blood mononucleated cells (PBMCs), bone marrow polychromatic erythrocytes of Balb/c mice, and human transformed cell lines derived from both gastric cancer (AGS cells) and leukaemia (K562 cells). Treatment with 6-PSA (30-150 µM) reduced the viability of AGS and K562 cells together with a moderate, but significant, increase in the frequency of micronucleated cells and the induction of DNA breakage (Comet Assay). Moreover, 6-PSA increased the apoptosis rate in both the AGS and K562 cell lines in a caspase 8-dependent manner. In contrast, neither cytotoxicity nor genotoxicity were observed in PBMCs or bone marrow polychromatic erythrocytes of Balb/c mice after treatment with low doses of 6-PSA (0.2-2.0 mg/Kg). Instead, 6-PSA treatment resulted in the inhibition of PBMC proliferation, which was reversible after the compound was removed. Additionally, 6-PSA treatments (2-20 mg/Kg) increased the frequency of mature polychromatic erythrocytes in the bone marrow, suggesting a possible effect on the differentiation process of immune cells. The present results indicate that 6-PSA induces cytotoxicity and moderate genotoxicity, together with an increase in the apoptosis rate, in a caspase 8-dependent manner in gastric cancer cells. In contrast, a low toxicity was observed when PBMCs were exposed to 6-PSA.
Assuntos
Ácidos Anacárdicos/toxicidade , Citostáticos/toxicidade , Mucosa Gástrica/efeitos dos fármacos , Leucócitos Mononucleares/efeitos dos fármacos , Mutagênicos/toxicidade , Animais , Apoptose/efeitos dos fármacos , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Caspase 8/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Transformada , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Feminino , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Humanos , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Desnaturação de Ácido Nucleico/efeitos dos fármacosRESUMO
A first approach is presented to explain the entropic structural transition observed in the persistence length of DNA complexes formed with intercalating drug molecules. The proposed model is based on calculating the effective persistence length of two entropic springs associated in series, one intercalated with drug molecules and the other without drugs. As the total drug concentration in the sample increases, the lengths of the two entropic springs vary, modifying the effective persistence length. The theoretical predictions of this model are then compared to experimental results, and a good accordance was obtained.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Bacteriófagos/genética , DNA Viral/química , DNA Viral/efeitos dos fármacos , Daunorrubicina/farmacologia , Etídio/farmacologia , Substâncias Intercalantes/farmacologia , Antibióticos Antineoplásicos/química , Simulação por Computador , Daunorrubicina/química , Elasticidade , Entropia , Etídio/química , Substâncias Intercalantes/química , Modelos Químicos , Conformação de Ácido Nucleico/efeitos dos fármacos , Desnaturação de Ácido Nucleico/efeitos dos fármacos , Pinças ÓpticasRESUMO
Synchronous fluorescence spectra measured in a flow-injection system with double pH gradient modulation constitute a new second-order signal which is herein studied for the quantitative determination of three fluoroquinolone antibiotics in spiked human urine samples. Because calibration is done using aqueous solutions of each of the three analytes ciprofloxacin, norfloxacin and ofloxacin, the fluorescent urine background makes it necessary to achieve the second-order advantage. Several second-order multivariate calibration algorithms were evaluated for this purpose: parallel factor analysis, unfolded and multiway partial least-squares with residual bilinearization, and multivariate curve resolution-alternating least-squares. The best analytical figures of merit, a root mean square error of 4-6 mg L(-1) (corresponding to a relative error of 4-6% for a calibration range from 0 to 200 mg L(-1) for each analyte), and a limit of detection of 4 mg L(-1) were obtained using partial least-squares (in the specific unfolded version) combined with residual bilinearization. Reasons for the improved success of this latter technique are provided on the basis of the analysis of simulated second-order data.