Relationship between oxidative stress and mitochondrial function in the post-conditioned heart

The pathways activated by post-conditioning may converge on the mitochondria, in particular on the mitochondrial permeability transition pore. We sought to characterize the inhibition status of the mitochondrial permeability transition early after the post-conditioning maneuver and before long reperfusion was established. We observed that post-conditioning maneuvers applied to isolated rat hearts, after a prolonged ischemia and before reperfusion, promoted cardiac mechanical function recovery and maintained mitochondrial integrity. These effects were evaluated by mitochondrial swelling, calcium transport, and NAD⁺ content measurements; the improvements were established before restoring a long lasting reperfusion period. Mitochondrial integrity was associated with a diminution in oxidative stress, since carbonylation of proteins was prevented and aconitase activity was preserved in the post-conditioned hearts, implying that ROS might mediate mitochondrial dysfunction and mPTP opening. In addition, we found that cytochrome release was significantly abolished in the post-conditioned heart, in contrast with conventionally reperfused hearts.     

New Pd(II) and Pt(II) complexes with N,S-chelated pyrazolonate ligands: molecular and supramolecular structure and preliminary study of their in vitro antitumoral activity

New Pd(II) and Pt(II) complexes [ML2] (HL=a substituted 2,5-dihydro-5-oxo-1H-pyrazolone-1-carbothioamide) have been synthesized by reacting K2MCl4 (M=Pd, Pt) or Pd(OAc)2 with beta-ketoester thiosemicarbazones. The structures of seven of these complexes were determined by X-ray diffraction. Although all exhibit a distorted square-planar coordination with trans- or (in one case) cis-[MN2S2] kernels, their supramolecular arrangements vary widely from isolated molecules to 3D-networks. The in vitro antitumoral assays performed with two HL ligands and their metal complexes showed significant cytostatic activity for the latter, with the most active [ML2] derivative (a palladium complex) being about sixteen times more active than cis-DDP against the cisplatinum-resistant cell line A2780cisR.     

Four glycoside hydrolases are differentially modulated by auxins, cytokinins, abscisic acid and gibberellic acid in apple fruit callus cultures

α-l-Arabinofuranosidase, α- and β-d-xylosidase, and β-d-glucosidase activity was detected in the soluble fraction (S-F) extracted with water and in the NaCl-released fraction (NaCl-F) extracted with a high-salt concentration buffer from apple callus cultures. The activity was found to be differentially modulated by the addition of various plant growth regulators (PGRs) to calluses that had lost their requirement for specific PGRs (“habituation” phenomenon). α-l-Arabinofuranosidase activity was 93%, 130%, 126% and 186% higher in the NaCl-F from IAA-, IBA-, ABA- and GA₃-treated callus than in that extracted from untreated callus while S-F α-l-arabinofuranosidase activity was only 71%, 24%, 55% and 66% higher, respectively. α-d-Xylosidase displayed low activity levels in both S-F and NaCl-F but 2iP-treated callus showed higher α-d-xylosidase activity in both fractions than the control. 2,4-D increased α-d-xylosidase activity by 110% in the NaCl-F but decreased it by 40% in the S-F. β-d-Xylosidase activity increased by 99% in S-F from 2iP-treated callus but slightly decreased in the NaCl-F. In GA₃-treated callus, NaCl-F β-d-xylosidase activity increased by 188%. S-F and NaCl-F from Picloram-treated callus showed undetectable or only slightly noticeable α-l-arabinofuranosidase, α-d-xylosidase and β-d-xylosidase activity. Interestingly, β-d-glucosidase activity rose 28-fold in the S-F extracted from Picloram-treated callus. β-d-glucosidase was the only enzyme assayed that greatly increased its NaCl-F activity after 10 subcultures, and the addition of any PGR to the callus culture –except for Picloram and ABA– decreased its activity, suggesting that this enzyme may be associated with certain stress conditions, such as PGR starvation or Picloram addition. This is the first report on glycoside hydrolases from fruit callus as modulated by different PGRs.     

Efficacy of Heterorhabditis baujardi LPP7 (Nematoda: Rhabditida) applied in Galleria mellonella (Lepidoptera: Pyralidae) insect cadavers to Conotrachelus psidii, (Coleoptera: Curculionidae) larvae

The guava weevil, Conotrachelus psidii, is a major pest of guava in Brazil causing severe reduction in fruit quality. We assessed its susceptibility to Heterhorhabditis baujardi LPP7 infective juveniles (IJs) in the greenhouse and under field conditions applying the nematodes in cadavers of seventh instar Galleria mellonella larvae. Field persistence of these nematodes in the soil was evaluated through G. mellonella-baiting. Insect cadaver concentrations of 2, 4 and 6 applied in pots in the greenhouse experiment caused significant mortality compared to the control. Significance differences were observed in the field between control and treatments only when six cadavers per 0.25 m² were applied. Infective juveniles from the cadavers persisted 6 weeks after application in the field, but decreased greatly thereafter. Our work demonstrates that H. baujardi LPP7 IJs emerging from G. mellonella cadavers can be efficacious against guava weevil fourth instar larvae. Also, we demonstrated the long-term persistence of IJs in the soil.     

Functional and structural roles of conserved cysteine residues in the carboxyl-terminal domain of the follicle-stimulating hormone receptor in human embryonic kidney 293 cells

The carboxyl-terminal segment of G protein-coupled receptors has one or more conserved cysteine residues that are potential sites for palmitoylation. This posttranslational modification contributes to membrane association, internalization, and membrane targeting of proteins. In contrast to other members of the glycoprotein hormone receptor family (the LH and thyroid-stimulating hormone receptors), it is not known whether the follicle-stimulating hormone receptor (FSHR) is palmitoylated and what are the effects of abolishing its potential palmitoylation sites. In the present study, a functional analysis of the FSHR carboxyl-terminal segment cysteine residues was carried out. We constructed a series of mutant FSHRs by substituting cysteine residues with alanine, serine, or threonine individually and together at positions 629 and 655 (conserved cysteines) and 627 (nonconserved). The results showed that all three cysteine residues are palmitoylated but that only modification at Cys629 is functionally relevant. The lack of palmitoylation does not appear to greatly impair coupling to G(s) but, when absent at position 629, does significantly impair cell surface membrane expression of the partially palmitoylated receptor. All FSHR Cys mutants were capable of binding agonist with the same affinity as the wild-type receptor and internalizing on agonist stimulation. Molecular dynamics simulations at a time scale of approximately 100 nsec revealed that replacement of Cys629 resulted in structures that differed significantly from that of the wild-type receptor. Thus, deviations from wild-type conformation may potentially contribute to the severe impairment in plasma membrane expression and the modest effects on signaling exhibited by the receptors modified in this particular position.     

Identification and characterization of Myosin from rat testicular peritubular myoid cells

In the mammalian testis, peritubular myoid cells (PMCs) surround seminiferous tubules. These cells are contractile, express the cytoskeletal markers of true smooth muscle-alpha-isoactin and F-actin-and participate in the contraction of seminiferous tubules during the transport of spermatozoa and testicular fluid to the rete testis. Myosin from PMCs (PMC-myosin) was isolated from adult rat testis and purified by cycles of assembly-disassembly and sucrose gradient centrifugation. PMC-myosin was recognized by a monoclonal anti-smooth muscle myosin antibody, and the peptide sequence shared partial homology with rat smooth muscle myosin-II, MYH11 (also known as SMM-II). Most PMC-myosin (95%) was soluble in the PMC cytosol, and purified PMC-myosin did not assemble into filaments in the in vitro salt dialysis assay at 4 degrees C, but did at 20 degrees C. PMC-myosin filaments are stable to ionic strength to the same degree as gizzard MYH11 filaments, but PMC-myosin filaments were more unstable in the presence of ATP. When PMCs were induced to contract by endothelin 1, a fraction of the PMC-myosin was found to be involved in the contraction. From these results we infer that PMCs express an isoform of smooth muscle myosin-II that is characterized by solubility at physiological ionic strength, a requirement for high temperature to assemble into filaments in vitro, and instability at low ATP concentrations. PMC-myosin is part of the PMC contraction apparatus when PMCs are stimulated with endothelin 1.     

Sexual dimorphism in the andromonoecious Euphorbia nicaeensis: effects of gender and inflorescence development

BACKGROUND AND AIMS: In andromonoecious taxa with separate floral types along the inflorescence, architectural or plastic effects can simulate floral sexual dimorphism. Both the primary and secondary sexual characteristics of the cyathia of the protogynous andromonoecious species Euphorbia nicaeensis were analysed according to their sex and arrangement on the inflorescence. METHODS: The production of male and hermaphrodite cyathia at each inflorescence level was surveyed in two natural populations. The longevity, size, pollen production and viability, and nectar secretion of both types of cyathia were checked between inflorescence levels and between sexes at the only level at which they occur together. This sampling method makes it possible to know whether differences between cyathia types are based on sex or are attributable to inflorescence development. KEY RESULTS: Male cyathia were produced predominantly at the first and second inflorescence levels, whereas at levels 3-5, the cyathia were almost exclusively hermaphrodite. Viable pollen production by male cyathia at the second inflorescence level was higher than that of hermaphrodite cyathia at the third level but, when males and hermaphrodites at the same level were compared, their pollen production was similar. Male and hermaphrodite cyathia were similar in size, irrespective of the inflorescence level, although the exclusively hermaphrodite cyathia of the last level were smaller. Both cyathium types produced similar amounts of sugar. However, male cyathia produced nectar during their whole lifespans, whereas hermaphrodites produced it exclusively during their male phase. Moreover, the nectary activity of male cyathia started earlier in the day than that of hermaphrodites. CONCLUSIONS: An apparent floral dimorphism exists in the primary sexual characteristics of Euphorbia nicaeensis because differences in pollen production between cyathium types are due to theirs positions. Similarly, differences affecting most secondary sexual characteristics are only apparent between the two cyathium types. However, E. nicaeensis shows a true but slight floral dimorphism in some of the secondary sex characters related to nectar secretion. The lack of nectar production by the female phase of the hermaphrodite cyathia of E. nicaeensis indicates that this is a deceit-pollinated species.