Copper(II) interactions with non-steroidal anti-inflammatory agents. III--3-Methoxyanthranilic acid as a potential *OH-inactivating ligand: a quantitative investigation of its copper handling role in vivo

Pharmacological activities of copper(II) complexes are a direct function of the nature of their ligands associated with the metal ion in vivo. Some of these, defined as *OH-inactivating ligands (G. Berthon, Agents Actions 39 (1993) 210-217), may act as specific "lures" for hydroxyl radicals at inflammatory sites and behave as pseudo-catalase-like agents. This property has been advanced for anthranilic acid (H. Miche, V. Brumas, G. Berthon, J. Inorg. Biochem. 68 (1997) 27-38). With a view to improve the chemical features required to render such inactive substances effective anti-inflammatory drugs through their association with copper(II), an in vitro investigation into copper(II) interactions with the anionic form of an anthranilic acid derivative, namely 3-methoxyanthranilate (Man), has been performed under experimental conditions pertaining in vivo. Copper(II)-Man complex equilibria have been determined using glass electrode potentiometry, then checked by UV-vis and mass spectrometries. Given the prime role of histidine as a copper(II) ligand in blood plasma, copper(II)-histidine-Man ternary equilibria have also been studied. Subsequent computer simulations of the distribution of copper(II) in the extracellular fluid revealed that Man can specifically mobilize Cu(II) ions under inflammatory conditions without affecting their distribution under normal physiological conditions. Thiobarbituric acid reactive substances (TBARS) tests conducted with respect to standardized copper-mediated Fenton-type reactions (P. Maestre, L. Lambs, J.P. Thouvenot, G. Berthon, Free Rad. Res. 20 (1994) 205-218) have shown that, like anthranilic acid, Man can effectively both increase the Fenton-like reactivity of copper and decrease the amount of TBARS detected in solution, i.e., act as a potential *OH-inactivating ligand.     

Genomic context analysis in Archaea suggests previously unrecognized links between DNA replication and translation

Background  

Comparative analysis of genomes is valuable to explore evolution of genomes, deduce gene functions, or predict functional linking between proteins. Here, we have systematically analyzed the genomic environment of all known DNA replication genes in 27 archaeal genomes to infer new connections for DNA replication proteins from conserved genomic associations.     

Copper--ligand interactions and the physiological free radical processes. Part 3. Influence of histidine, salicylic acid and anthranilic acid on copper-driven Fenton chemistry in vitro

With a view to the possible use of copper(II)-^·OH inactivating ligand (OIL) complexes as regulators of inflammation, the reactivity of the copper(II)-ascorbate system with hydrogen peroxide has been investigated in the presence of three key substances: histidine (the main copper(II) low molecular mass ligand in extracellular fluid), salicylic acid (the well-known non-steroidal antiinflammatory drug, previously shown to be potentiated by copper(II) in animal models of inflammation), and anthranilic acid (an inactive substance by itself, known to be activated by copper(II) in the same models) at physiological pH (7.4) and inflammatory pH (5.5).

Such substances may affect the amount of TBARS detected in solution following copper-mediated Fenton-like reactions through three distinct mechanisms: (i) by decreasing the Cu(II)/Cu(I) redox potential, i.e. at the expense of ^·OH radical production, (ii) by scavenging ^·OH radicals in the body of the solution, and/or (iii) by acting as a true OIL, i.e. at the expense of ^·OH detection. Redox potential measurements of initial solutions have been performed in parallel to TBARS determinations to help discriminate between different ligand influences. Computer-aided speciation has been used to understand the role of copper(II) distribution on the ligand effects characterised.

Contrary to previous interpretations, histidine has been found to mainly affect ^·OH production by lowering the redox potential of the Cu(II)/Cu(I) couple. Salicylate, which has no effect on ^·OH production, has been confirmed to mainly scavenge ^·OH radicals in the body of the solution. Anthranilate, which both increases ^·OH production and decreases ^·OH detection, behaves as a potential OIL.

These results tend to confirm our previous hypothesis that copper potentiation of antiinflammatory substances is indirect, i.e. independent of any interaction between metal and drug, whereas copper activation of substances that are inactive by themselves results from specific metal-substance interactions taking place at inflammatory sites.     

Aluminum speciation studies in biological fluids. Part 6. Quantitative investigation of aluminum(III)-tartrate complex equilibria and their potential implications for aluminum metabolism and toxicity

Recent epidemiological studies have confirmed the existence of a correlation between aluminum level in low-silica drinking water and prevalence of Alzheimer's disease. Also, oral aluminum-based phosphate binders and antacids may induce acute aluminum toxicity. Whatever the source of the metal ingested, its bioavailability is a function of the chemical forms under which it occurs in the gastrointestinal tract, i.e. of the ligands with which the Al3+ ion may associate. Dietary acids in particular can favor the bioavailability of aluminum in different ways: by increasing its solubility, by complexing it into neutral species, and/or by acting indirectly on its absorption process. Among these, tartaric acid is commonly found in fruits and in industrial foods and drinks, and may therefore be ingested together with environmental or/and therapeutic aluminum. The present work examines its potential influence on aluminum bioavailability. Firstly, Al(III)-tartrate complex formation constants have been determined under physiological conditions (37 degrees C, 0.15 M NaCl). Then these constants have been used to simulate the influence of tartrate on aluminum speciation in different gastrointestinal situations in which phosphate was also taken into account. Under normal conditions of aluminum contamination, tartrate is expected to keep the metal soluble throughout the whole pH range of the small intestine, which is likely to enhance its bioavailability. Even at low concentrations, tartrate also gives rise to two neutral complexes that span over the 1.5-7.5 pH interval, a phenomenon that is aggravated by increased aluminum levels as may result from aluminum hydroxide therapy. The co-occurrence of dietary phosphate reduces the fraction of aluminum neutralized by tartrate under normal conditions, but this effect quickly decreases with increasing aluminum doses. Even the therapeutic use of aluminum phosphate is not expected to be totally safe in the presence of tartaric acid. As plasma simulations show that no aluminum mobilization can be expected from tartrate that could enhance aluminum excretion, avoiding ingestion of tartaric acid during any form of aluminum-based therapy appears advisable.     

Defining pallial and subpallial divisions in the developing Xenopus forebrain

To shed light on the organisation of the Xenopus laevis telencephalon, we have used two sets of developmental regulators: genes acting in early regional specification (x-Dll3, x-Nkx2.1, x-Emx1, x-Pax6, x-Eomes) or in cell determination (x-Lhx5 and x-Lhx7). After expression patterns analysis, separately or combined, on whole-mount brains and serial sections, we identify the Xenopus pallium and subpallium, and the subdivisions herein. The data show a conservation of the same basic Bauplan for Xenopus forebrain patterning compared to other vertebrates, and suggest the possibility for LIM-homeodomain genes to be candidate downstream target of the regionalization genes. Comparing the relative sizes of the deduced subdivisions, Xenopus seems to have an intermediate phylogenetic position in terms of pallium contribution to the telencephalon, and ventral pallium contribution to the pallium.     

Ethnopharmacology and bioinformatic combination for leads discovery: application to phospholipase A(2) inhibitors.

A program combining ethnopharmacology and bioinformatic approaches has successfully been applied on anti-inflammatory activity. (i) An ethnobotanical study allowed the identification of several plants associated with putative anti-inflammatory properties as potential leads. (ii) On the other hand, it is well known that phospholipase A(2) is a target implicated in the pro-inflammatory process. Thus, (iii) some selected plant extracts were experimentally tested on phospholipase A(2). Finally, (iv) these experimental results combined with bioinformatic tools, such as database exploitation and molecular modeling, allowed to suggest that one compound, betulin and its oxidative form betulinic acid, might be responsible of the anti-PLA(2) activity. This suggestion was confirmed experimentally.     

Metal ion-tetracycline interactions in biological fluids. 2. Potentiometric study of magnesium complexes with tetracycline, oxytetracycline, doxycycline, and minocycline, and discussion of their possible influence on the bioavailability of these antibiotics in blood plasma

The formation constants of the various complexes formed by magnesium with four tetracycline derivatives, namely, tetracycline itself, oxytetracycline, doxycycline, and minocycline, were determined by potentiometry over large pH ranges under experimental conditions pertaining to blood plasma (37 degrees C, NaCl 0.15 mol dm-3). The results were used, together with those previously obtained on the complexation of these tetracyclines with proton and calcium, to assess the influence of the two alkali earth metal ions on the bioavailability of these drugs in blood plasma. Accordingly, simulations of the distribution of the four tetracyclines into their different proton and metal complex species were calculated. The distributions confirm that, in combination with the protein-bound fraction of the tetracyclines, the metal-bound fraction represents more than 99% of these drugs in plasma, the extent of their free fraction commonly being less than 1%.     

Shivering thermogenesis in the neonatal pig

(I) Shivering intensity and metabolic rate were determined in Large White pigs aged 2, 24, 48 h and 5 d, at temperatures ranging from thermoneutrality (36°C) to cold (20°C). (2) Shivering is the main heat producing mechanism, the absence of nonshivering thermogenesis being implied by both the absence of delay between the onset of shivering (S_tt) and the increase in metabolic rate (L_ct) and by the linearity of the relationship between metabolic rate and shivering intensity in the cold. (3) For a comparable thermal demand, shivering intensity decreased with age whereas cold induced heat production remained constant, which suggests that the thermogenic efficiency of shivering is improved during the first 5 days of life.     

Aluminum speciation studies in biological fluids. Part 5. A quantitative investigation of A1(III) complex equilibria with desferrioxamine, 2,3-dihydroxybenzoic acid, Tiron, CP20 (L1), and CP94 under physiological conditions, and computer-aided assessment of the aluminum-mobilizing capacities of these ligands in vivo.

While the involvement of environmental aluminum toxicity in the advent of senile dementias is still debated, acute aluminum toxicity of iatrogenic origin is well documented. So far, the only treatment available against it has been desferrioxamine (DFO), which induces major side effects. New drugs are thus highly desirable, and possible DFO substitutes have already been considered through various techniques. An important test for such new drugs is to assess their A1-mobilizing capacity in vivo. This can be done by computer-aided speciation provided formation constants for the corresponding A1(III) complexes are known beforehand. The present work reports an investigation of A1(III) complex equilibria with five sequestering ligands including DFO, and predicts the respective capacities of these to mobilize aluminum in vivo under normal and inflammatory conditions.