On the interactions involving serine proteases obtained from Monacrosporium thaumasium (Ascomycota: Orbiliomycetes) and deoxyribonucleic acid (DNA): biological macromolecules in action.

The use of force spectroscopy approaches performed with optical tweezers can be very useful in determining the binding modes and the physical chemistry of DNA interactions with ligands, from small drugs to proteins. Helminthophagous fungi, on the other hand, have important enzyme secretion mechanisms for various purposes, and the interactions between such enzymes and nucleic acids are very poorly studied. Therefore, the main goal of the present work was to investigate, at the molecular level, the mechanisms of interaction between fungal serine proteases and the double-stranded (ds) DNA molecule. Experimental assays performed with this single molecule technique consist in exposing different concentrations of the protease of this fungus to dsDNA until saturation while monitoring the changes on the mechanical properties of the macromolecular complexes formed, from where the physical chemistry of the interaction can be deduced. It was found that the protease binds strongly to the double-helix, forming aggregates and changing the persistence length of the DNA molecule. The present work thus allowed us to infer information at the molecular level on the pathogenicity of these proteins, an important class of biological macromolecules, when applied to a target specimen.

Antinociceptive effect of PnTx4(5-5), a peptide from Phoneutria nigriventer spider venom, in rat models and the involvement of glutamatergic system.

BACKGROUND: The venom of Phoneutria nigriventer spider is a source of numerous bioactive substances, including some toxins active in insects. An example is PnTx4(5-5) that shows a high insecticidal activity and no apparent toxicity to mice, although it inhibited NMDA-evoked currents in rat hippocampal neurons. In this work the analgesic activity of PnTx4(5-5) (renamed Γ-ctenitoxin-Pn1a) was investigated. METHODS: The antinociceptive activity was evaluated using the paw pressure test in rats, after hyperalgesia induction with intraplantar injection of carrageenan or prostaglandin E2 (PGE2). RESULTS: PnTx4(5-5), subcutaneously injected, was able to reduce the hyperalgesia induced by PGE2 in rat paw, demonstrating a systemic effect. PnTx4(5-5) administered in the plantar surface of the paw caused a peripheral and dose-dependent antinociceptive effect on hyperalgesia induced by carrageenan or PGE2. The hyperalgesic effect observed in these two pain models was completely reversed with 5 µg of PnTx4(5-5). Intraplantar administration of L-glutamate induced hyperalgesic effect that was significantly reverted by 5 µg of PnTx4(5-5) injection in rat paw. CONCLUSION: The antinociceptive effect for PnTx4(5-5) was demonstrated against different rat pain models, i.e. induced by PGE2, carrageenan or glutamate. We suggest that the antinociceptive effect of PnTx4(5-5) may be related to an inhibitory activity on the glutamatergic system.

Cardiac protein changes in rats after soybean oil treatment: a proteomic study.

BACKGROUND: Several studies show that the consumption of vegetable oils, such as soybean oil, rich in polyunsaturated fatty acids (PUFAs) has beneficial health effects by preventing or reducing the risk factors of cardiovascular diseases. While the demonstration of beneficial effects of the consumption of unsaturated fatty acids on the cardiovascular system has been proven in a macroscopic level, the molecular/cellular mechanisms responsible for this phenomenon are poorly understood. METHODS: In this work, a comparative proteomic approach, two-dimensional gel electrophoresis (2-DE) coupled to mass spectrometry (MALDI-TOF/TOF), was applied to investigate proteome differences in the left ventricle (LV) of rats that received 0.1 mL of soybean oil intramuscularly for 15 days (treated group-TR) and rats that had not (control group-CT). RESULTS: Soybean oil treatment improved left ventricular function, TR animals presented lower value of LVEDP and significantly changed LV proteome. The protein profile of VE revealed differences in the expression of 60 protein spots (p<0.05) between the experimental groups (CT and TR), 14 of those were identified by MS and MS/MS, and 12 of the 14 being non-redundant proteins. Robust changes were detected in proteins involved in cellular structure and antioxidant system and muscular contraction. CONCLUSIONS: The TR group presented an increase in the intensity of proteins involved in muscle contraction (myosin light chain-3 (3-MCL), creatine kinase M (CKM)) and thireodoxin, an antioxidant enzyme. Low intensity cytoskeletal protein, desmin, was also detected in TR animals. The results suggest that soybean oil induces changes in the levels of heart proteins which may partially account for the underlying mechanisms involved in the benefits provided by oils rich in polyunsaturated fatty acids.

Effects of captopril on cardiovascular reflexes and respiratory mechanisms in rats submitted to monocrotaline-induced pulmonary arterial hypertension.

BACKGROUND: Pulmonary Arterial Hypertension (PAH) is a disease associated with increased arteriolar resistance in the lungs. Due to hypoxemia, some physiological mechanisms can be posteriorly affected, including respiratory and cardiovascular reflexes, but this has not yet been fully investigated. This study aimed to evaluate how these mechanisms were affected by monocrotaline (MCT)-induced PAH and the possible therapeutic role of angiotensin converting enzyme inhibitor (ACEi), captopril, in reversing this remodeling process. METHODS AND RESULTS: Groups of Wistar rats received MCT injections (60 mg kg(-1)). Three weeks later, they received captopril (CPT, 100 mg kg(-1)) in their drinking water (MCT + CPT) or water alone (MCT) for 2 weeks. As control, saline-treated animals received captopril in their drinking water (CPT) or water alone (CON), also for 2 weeks. Results showed that PAH was fully induced in the MCT group, evidenced by a high pulmonary index. Gasometrical and respiratory analyses showed hypoxemia and compensatory hyperventilation. CPT treatment brought these parameters to similar values to those observed in the CON group. We observed that autonomic dysfunction in the MCT group was suppressed by CPT. Finally, cardiovascular reflexes analysis showed increased chemoreflex responses in the MCT group, while baroreflex sensibility was decreased. Surprisingly, CPT normalized these reflex responses to values similar to the CON group. CONCLUSIONS: The present study demonstrates that MCT-induced PAH induces compensatory respiratory responses, dysautonomia, and baroreflex dysfunction and increases chemoreflex responses. The data also indicate that CPT was effective in reversing these cardio-respiratory disorders, suggesting that ACEi could be a potential therapeutic target for PAH.