Synthesis and cardiovascular activity of metoprolol analogues

The synthesis of four novel analogues of metoprolol, a well-known beta1-blocker used to reduce arterial blood pressure, is described. The preparation of (2S,2'S)-7, (2R,2'S)-7, (2R,2'R)-8, and (2S,2'R)-8 was based on the reaction of racemic 2-[4-(2'-methoxyethyl)-phenoxymethyl]-oxirane (4) with (R)- or (S)-2-amino-1-butanol. Salient characteristics of analogues 7 and 8 relative to metoprolol are the incorporation of an additional stereogenic center, as well as a methyl group and a hydroxyl function on the nitrogen-containing chain. These novel derivatives present significant hypotensive and bradycardiac activity, although no blocking action toward beta1 and beta2 adrenergic receptor.     

Betaine improves growth, but does not induce whole body or hepatic palmitate oxidation in swine (Sus scrofa domestica)

Dietary betaine may reduce carcass fat in growing pigs. We explored the effects of betaine on short-term growth and in vivo and in vitro fatty acid oxidation. Pigs were housed in metabolism crates and fed diets containing either 0% (control), 0.125% or 0.5% betaine at 80% of ad libitum energy intake. Fatty acid oxidation was measured during intravenous infusions of 1-(13)C-palmitate and in hepatocytes incubated in the presence or absence of betaine and carnitine. CO2 and palmitate isotopic enrichments were determined by mass spectrometry. Pigs consuming 0.125% and 0.5% betaine for at least 9 days had growth rates that were 38% and 12% greater than controls, respectively. Feed efficiency was also improved with betaine. Fasting increased palmitate oxidation rates 7-8-fold (P < 0.01), but betaine had no effect in either the fed or fasted state (P > 0.1). For hepatocytes, carnitine but not betaine enhanced palmitate oxidation. This response suggests that previously observed reduction in adipose accretion must be via a mechanism other than oxidation. Betaine had no effect on plasma non-esterified fatty acids or urea nitrogen. Under the confinement conditions in this study, dietary betaine improved animal growth responses, but it had no apparent effect on either whole body or hepatic fatty acid oxidation.     

Proteomic identification of brain proteins that interact with dynein light chain LC8

Cytoplasmic dynein is a large minus end-directed microtubule motor that translocates cargos towards the minus end of microtubules. Light chain 8 of the dynein machinery (LC8) has been reported to interact with a large variety of proteins that possess K/RSTQT or GIQVD motifs in their sequence, hence permitting their transport in a retrograde manner. Yeast two-hybrid analysis has revealed that in brain, LC8 associates directly with several proteins such as neuronal nitric oxide synthase, guanylate kinase domain-associated protein and gephyrin. In this work, we report the identification of over 40 polypeptides, by means of a proteomic approach, that interact with LC8 either directly or indirectly. Many of the neuronal proteins that we identified cluster at the post-synaptic terminal, and some of them such as phosphofructokinase, lactate dehydrogenase or aldolase are directly involved in glutamate metabolism. Other pool of proteins identified displayed the LC8 consensus binding motif. Finally, recombinant LC8 was produced and a library of overlapping dodecapeptides (pepscan) was employed to map the LC8 binding site of some of the proteins that were previously identified using the proteomic approach, hence confirming binding to the consensus binding sites.     

Thermal degradation of allicin in garlic extracts and its implication on the inhibition of the in-vitro growth of Helicobacter pylori

Allicin, the main active principle related to Allium sativum chemistry, is considered to be responsible for the bacteriostatic properties of garlic. The work described here has demonstrated the direct implication of the allicin present in solvent-free garlic extracts obtained with ethanol (ethanolic garlic extract, EGE) and acetone (acetonic garlic extract, AGE) in the inhibition of the in-vitro growth of Helicobacter pylori (Hp), the bacterium responsible for serious gastric diseases such as ulcers and even gastric cancer. The evolution of allicin concentration as a function of time and temperature has been the subject of a kinetic study. The reaction order, activation energy, and preexponential factor (in accordance with Arrhenius theory) have been determined for the decomposition process of allicin in these organic media. First-order decomposition, an activation energy of 97.4 kJ/mol, and an Arrhenius preexponential factor of 8.9 x 10(10) s(-1) have been determined for allicin in EGE. For allicin in AGE the kinetic order determined was 1.5, the activation energy 184.5 kJ/mol, and the preexponential factor 3.1 x 10(24) s(-1) (mg/L)(-0.5). The presence or absence of allicin in these garlic products was found to be crucial for the inhibition of the in-vitro growth of Hp, as demonstrated by microbiological analysis for AGE. A relationship has been identified between the effectiveness and durability of the anti-Hp properties shown by AGE and the allicin content of these products. The bacteriostatic properties were active for up to 10 months if the samples were maintained at 6 degrees C.     

Benzimidazole derivatives. Part 5: design and synthesis of new benzimidazole-arylpiperazine derivatives acting as mixed 5-HT1A/5-HT3 ligands

A series of new mixed benzimidazole-arylpiperazine derivatives were designed by incorporating in general structure III the pharmacophoric elements of 5-HT(1A) and 5-HT(3) receptors. Compounds 1-11 were synthesized and evaluated for binding affinity at both serotoninergic receptors, all of them exhibiting high 5-HT(3)R affinity (K(i)=10-62nM), and derivatives with an o-alkoxy group in the arylpiperazine ring showing nanomolar affinity for the 5-HT(1A)R (K(i)=18-150nM). Additionally, all the synthesized compounds were selective over alpha(1)-adrenergic and dopamine D(2) receptors (K(i)>1000-10,000nM). Compound 3 was selected for further pharmacological characterization due to its interesting binding profile as mixed 5-HT(1A)/5-HT(3) ligand with high affinity for both receptors (5-HT(1A): K(i)=18.0nM, 5-HT(3): K(i)=27.2nM). In vitro and in vivo findings suggest that this compound acts as a partial agonist at 5-HT(1A)Rs and as a 5-HT(3)R antagonist. This novel mixed 5-HT(1A)/5-HT(3) ligand was also effective in preventing the cognitive deficits induced by muscarinic receptor blockade in a passive avoidance learning test, suggesting a potential interest in the treatment of cognitive dysfunction.     

Interleukin-1 beta (IL-1beta) induces tumor necrosis factor alpha (TNF-alpha) expression on mouse myeloid multipotent cell line 32D cl3 and inhibits their proliferation

Interleukin-1 alpha (IL-1alpha) and beta (IL-1beta) are well known factors that stimulate hematopoiesis, nevertheless there are reports that show that they can also inhibit this activity. While both IL-1alpha and IL-1beta induce the expression of hematopoietic cytokines, such as growth factors and their receptors on myeloid cells, helping thus to regulate hematopoiesis, it is not known if their inhibitory activity is also mediated through the induction of other specific cytokines. In this work we show that recombinant human IL-1beta (rhIL-1beta) inhibits the proliferation of a mouse IL-3-dependent myeloid multipotent cell line (32D cl3), without inducing its differentiation. We show that rhIL-1beta induces in 32D cl3 cells the expression of the tumor necrosis factor alpha (TNF-alpha) gene, a well known growth inhibitor, and that the rhIL-1beta growth inhibition property on 32D cl3 cells is partially due to this secreted TNF-alpha, hinting thus that the inhibition of hematopoiesis by IL-1 is mediated through other induced cytokines.     

In vitro effects of LPS, IL-2, PDGF and CRF on haemocytes of Mytilus galloprovincialis Lmk

The cells in charge of the innate immune response in the marine mussel Mytilus galloprovincialis Lmk. are the haemocytes. These cells respond in different ways to agents such as lipopolysaccharide (LPS), interleukin-2 (IL-2), platelet-derived growth factor (PDGF) and corticotropin releasing factor (CRF). After stimulation of the haemocytes, the expression of molecules reactive with monoclonal antibodies raised to the alpha chain of the IL-2 receptor, present in their membrane, differed depending on the agent used. The same happened with regard to the levels of dopamine, adrenaline and noradrenaline released to the medium by the haemocytes. It should also be noted that no catecholamine release was detected and the level of expression of IL-2Ralpha showed no significant variation in cultured cells that had not been treated with inducers. These facts would indicate that most haemocytes were in the same starting condition at the moment that the stimulation was performed. Therefore, cultured haemocytes can be a highly reliable model in the study of the innate immune system.     

A novel locus for autosomal dominant nonsyndromic hearing loss (DFNA44) maps to chromosome 3q28–29

Hereditary non-syndromic sensorineural hearing loss (NSSHL) is a genetically highly heterogeneous group of disorders. Autosomal dominant forms account for up to 20% of cases. To date, 39 loci have been identified by linkage analysis of affected families that segregate NSSHL forms in the autosomal dominant mode (DFNA). Investigation of a large Spanish pedigree with autosomal dominant inheritance of bilateral and progressive NSSHL of postlingual onset excluded linkage to known DFNA loci and, in a subsequent genome-wide scan, the disorder locus was mapped to 3q28-29. A maximum two-point LOD score of 4.36 at theta=0 was obtained for marker D3S1601. Haplotype analysis placed the novel locus, DFNA44, within a 3-cM genetic interval defined by markers D3S1314 and D3S2418. Heteroduplex analysis and DNA sequencing of coding regions and exon/intron boundaries of two genes (CLDN16 and FGF12) in this interval did not reveal disease-causing mutations.     

Crystal structure of the vertebrate NADP(H)-dependent alcohol dehydrogenase (ADH8)

The amphibian enzyme ADH8, previously named class IV-like, is the only known vertebrate alcohol dehydrogenase (ADH) with specificity towards NADP(H). The three-dimensional structures of ADH8 and of the binary complex ADH8-NADP(+) have been now determined and refined to resolutions of 2.2A and 1.8A, respectively. The coenzyme and substrate specificity of ADH8, that has 50-65% sequence identity with vertebrate NAD(H)-dependent ADHs, suggest a role in aldehyde reduction probably as a retinal reductase. The large volume of the substrate-binding pocket can explain both the high catalytic efficiency of ADH8 with retinoids and the high K(m) value for ethanol. Preference of NADP(H) appears to be achieved by the presence in ADH8 of the triad Gly223-Thr224-His225 and the recruitment of conserved Lys228, which define a binding pocket for the terminal phosphate group of the cofactor. NADP(H) binds to ADH8 in an extended conformation that superimposes well with the NAD(H) molecules found in NAD(H)-dependent ADH complexes. No additional reshaping of the dinucleotide-binding site is observed which explains why NAD(H) can also be used as a cofactor by ADH8. The structural features support the classification of ADH8 as an independent ADH class.