An unusual glucoside from Cleistanthus gracilis

Extraction of roots and stems of Cleistanthus gracilis furnished common triterpenes, plant sterols and the unusual glucoside (+) gracicleistanthoside, the glucoside of 2-beta-hydroxy-8-azabicyclo-(5,2,0)-4beta,9beta-epoxynona-5,7-diene.     

Combined effect of vanadium(V) and chromium(III) on lipid peroxidation in liver and kidney of rats

Since chromium(III) was demonstrated to have antioxidative action, we have decided to study the effect of this element on V-induced LPO in liver and kidney of rats. Outbred 2-month-old, albino male Wistar rats received daily, for a period of 12 weeks: group I (control), deionized water to drink; group II, sodium metavanadate (SMV) solution at a concentration of 0.100mgV/mL; group III, chromium chloride (CC) solution at a concentration of 0.004mgCr/mL and group IV, SMV-CC solution at a concentration of 0.100mgV and 0.004mgCr/mL. The particular experimental groups took up with drinking water about 8.6mgV/kg b.w./24h (group II), 0.4mgCr/kg b.w./24h (group III), 9mgV and 0.36mgCr/kg b.w./24h (group IV). The V- or Cr-treated groups had higher concentrations of these two elements in liver and kidney compared to the controls. The administration of vanadium alone caused a significant decrease in fluid intake and in body weight gain compared to the controls. In liver supernatants obtained from all tested rats a statistically significant increase in MDA concentration was demonstrated in spontaneous LPO in comparison with the control rats. Moreover, in rats intoxicated with vanadium alone a statistically significant increase in liver MDA level was observed in the presence of 100microM NaVO(3). Instead, in supernatants of liver received from rats treated with chromium alone, a statistically significant increase in MDA concentration in comparison with the controls was found in the presence of 400microM NaVO(3). In kidney supernatants obtained from rats treated with chromium alone, a statistically significant increase in lipid peroxidation was shown in the presence of 30microM FeSO(4) and 400microM NaVO(3). These results show that the tested doses of vanadium(V) and chromium(III) ingested by rats with their drinking water caused significant alterations in internal organs, especially in liver. Under the conditions of our experiment, Cr(III) did not demonstrate antioxidant action, it rather had an oxidant effect.     

Resilience of Rhodobacter sphaeroides cytochrome bc1 to heme c1 ligation changes

Typically, c hemes are bound to the protein through two thioether bonds to cysteines and two axial ligands to the heme iron. In high-potential class I c-type cytochromes, these axial ligands are commonly His-Met. A change in this methionine axial ligand is often correlated with a dramatic drop in the heme redox potential and loss of function. Here we describe a bacterial cytochrome c with an unusual tolerance to the alternations in the heme ligation pattern. Substitution of the heme ligating methionine (M185) in cytochrome c1 of the Rhodobacter sphaeroides cytochrome bc1 complex with Lys and Leu lowers the redox midpoint potential but not enough to prevent physiologically competent electron transfer in these fully functional variants. Only when Met-185 is replaced with His is the drop in the redox potential sufficiently large to cause cytochrome bc1 electron transfer chain failure. Functional mutants preserve the structural integrity of the heme crevice: only the nonfunctional His variant allows carbon monoxide to bind to reduced heme, indicating a significant opening of the heme environment. This range of cytochrome c1 ligand mutants exposes both the relative resilience to sixth axial ligand change and the ultimate thermodynamic limits of operation of the cofactor chains in cytochrome bc1.     

Cellular uptake of S413-PV peptide occurs upon conformational changes induced by peptide-membrane interactions

In face of accumulated reports demonstrating that uptake of some cell-penetrating peptides occurs through previously described endocytic pathways, or is a consequence of cell fixation artifacts, we conducted a systematic analysis on the mechanism responsible for the cellular uptake of the S4(13)-PV karyophilic cell-penetrating peptide. The results reviewed here show that the S4(13)-PV peptide is able to very efficiently accumulate inside live cells in a rapid, non-toxic and dose-dependent manner, through a mechanism distinct from endocytosis. Comparative analysis of peptide uptake by mutant cells lacking heparan sulfate proteoglycans demonstrates that, although not mandatory, their presence at cell surface facilitates the cellular uptake of the S4(13)-PV peptide. Furthermore, we demonstrate that upon interaction with lipid vesicles, the S4(13)-PV peptide undergoes significant conformational changes that are consistent with the formation of helical structures. Such conformational changes occur concomitantly with a penetration of the peptide into the lipid bilayer, strongly suggesting that the resulting helical structures are crucial for the non-endocytic cellular uptake of the S4(13)-PV peptide. Overall, our data support that, rather than endocytosis, the cellular uptake of the S4(13)-PV cell-penetrating peptide is a consequence of its direct translocation through cell membranes following conformational changes induced by peptide-membrane interactions.     

The human placental derived BeWo cell line: a useful model for selecting Plasmodium falciparum CSA-binding parasites

Chondroitin sulfate A (CSA) present in the placental intervillous blood spaces has been described as the main receptor involved in the massive sequestration of Plasmodium falciparum parasitized erythrocytes to the placenta. Placental parasite isolates are functionally distinct from isolates that sequester in other organs, because they do not cytoadhere to CD36 but instead bind to CSA. To investigate for the parasites molecules associated with the CSA adhesion phenotype, different methodologies have been developed to select for CSA-binding lines in vitro mainly using non-placental sources of CSA that differ in their sulfation pattern. In this study, we show that the human trophoblastic BeWo cell line is a very efficient alternative to select for the CSA-binding phenotype in parasitized erythrocytes.     

Doubly uniparental inheritance is associated with high polymorphism for rearranged and recombinant control region haplotypes in Baltic Mytilus trossulus

Many bivalve species, including mussels of the genus Mytilus, are unusual in having two mtDNA genomes, one inherited maternally (the F genome) and the other inherited paternally (the M genome). The sequence differences between the genomes are usually great, indicating ancient divergence predating speciation events. However, in Mytilus trossulus from the Baltic, both genomes are similar to the F genome from the closely related M. edulis. This study analyzed the mtDNA control region structure in male and female Baltic M. trossulus mussels. We show that a great diversity of structural rearrangements is present in both sexes. Sperm samples are dominated by recombinant haplotypes with M. edulis M-like control region segments, some having large duplications. By contrast, the rearranged haplotypes that dominate in eggs lack segments from this M genome. The rearrangements can be explained by a combination of tandem duplication, deletion, and intermolecular recombination. An evolutionary pathway leading to the recombinant haplotypes is suggested. The data are also considered in relation to the hypothesis that the M. edulis M-like control region sequence is necessary to confer the paternal role on genomes that are otherwise F-like.     

EMG and MMG of agonist and antagonist muscles as a function of age and joint angle.

The aim of this study was to determine the effect of elbow joint position on electromyographic (EMG) and mechanomyographic (MMG) activities of agonist and antagonist muscles in young and old women. Surface EMG and MMG were recorded from the triceps and biceps brachii, and brachioradialis muscles during isometric elbow extensions in young and old women. The measurements were carried out at an optimal joint angle (A(o)), as well as at smaller (A(s) = A(o) - 30 degrees ) and larger (A(l) = A(o) + 30 degrees ) angles. The normalized to force EMG amplitude (RMS-EMG/F) was smaller in old women compared to young in all muscles. The RMS-EMG/F of the triceps brachii muscle was not affected by muscle length while that of the biceps brachii and brachioradialis muscles increased at shortest muscle length in both groups. The normalized to force MMG amplitude (RMS-MMG/F) was smaller in old than in young in the triceps brachii muscle only. There was an increase in RMS-MMG/F with triceps brachii and biceps brachii muscle shortening in both groups, and in the brachioradialis muscle -- in young only. Compared to young, older women exhibited a bigger force fluctuation during maximum voluntary contraction, but these did not contribute significantly to the RMS-MMG. Skinfold thickness accounted for the RMS-EMG/F and RMS-MMG/F differences seen between old and young women in the biceps brachii muscle only. It is concluded that, the EMG and MMG response to muscles length change in agonist and antagonist muscles is generally similar in old and young women but the optimal angle shifts toward a bigger value in older women.     

A novel approach to measure the contribution of matrix metalloproteinase in the overall net proteolytic activity present in synovial fluids of patients with arthritis

Despite decades of research, only a very limited number of matrix metalloproteinase (MMP) inhibitors have been successful in clinical trials of arthritis. One of the central problems associated with this failure may be our inability to monitor the local activity of proteases in the joints since the integrity of the extracellular matrix results from an equilibrium between noncovalent, 1:1 stoichiometric binding of protease inhibitors to the catalytic site of the activated forms of the enzymes. In the present work, we have measured by flow cytometry the net proteolytic activity in synovial fluids (SF) collected from 95 patients with osteoarthritis and various forms of inflammatory arthritis, including rheumatoid arthritis, spondyloarthropathies, and chronic juvenile arthritis. We found that SF of patients with inflammatory arthritis had significantly higher levels of proteolytic activity than those of osteoarthritis patients. Moreover, the overall activity in inflammatory arthritis patients correlated positively with the number of infiltrated leukocytes and the serum level of C-reactive protein. No such correlations were found in osteoarthritis patients. Members of the MMP family contributed significantly to the proteolytic activity found in SF. Small-molecular-weight MMP inhibitors were indeed effective for inhibiting proteolytic activity in SF, but their effectiveness varied greatly among patients. Interestingly, the contribution of MMPs decreased in patients with very high proteolytic activity, and this was due both to a molar excess of tissue inhibitor of MMP-1 and to an increased contribution of other proteolytic enzymes. These results emphasize the diversity of the MMPs involved in arthritis and, from a clinical perspective, suggest an interesting alternative for testing the potential of new protease inhibitors for the treatment of arthritis.     

Molecular modeling of hair keratin/peptide complex: Using MM-PBSA calculations to describe experimental binding results

Molecular dynamics simulations of a keratin/peptide complex have been conducted to predict the binding affinity of four different peptides toward human hair. Free energy calculations on the peptides' interaction with the keratin model demonstrated that electrostatic interactions are believed to be the main driving force stabilizing the complex. The molecular mechanics-Poisson-Boltzmann surface area methodology used for the free energy calculations demonstrated that the dielectric constant in the protein's interior plays a major role in the free energy calculations, and the only way to obtain accordance between the free energy calculations and the experimental binding results was to use the average dielectric constant.