Association between the ACCN1 gene and multiple sclerosis in Central East Sardinia

Multiple genome screens have been performed to identify regions in linkage or association with Multiple Sclerosis (MS, OMIM 126200), but little overlap has been found among them. This may be, in part, due to a low statistical power to detect small genetic effects and to genetic heterogeneity within and among the studied populations. Motivated by these considerations, we studied a very special population, namely that of Nuoro, Sardinia, Italy. This is an isolated, old, and genetically homogeneous population with high prevalence of MS. Our study sample includes both nuclear families and unrelated cases and controls. A multi-stage study design was adopted. In the first stage, microsatellites were typed in the 17q11.2 region, previously independently found to be in linkage with MS. One significant association was found at microsatellite D17S798. Next, a bioinformatic screening of the region surrounding this marker highlighted an interesting candidate MS susceptibility gene: the Amiloride-sensitive Cation Channel Neuronal 1 (ACCN1) gene. In the second stage of the study, we resequenced the exons and the 3' untranslated (UTR) region of ACCN1, and investigated the MS association of Single Nucleotide Polymorphisms (SNPs) identified in that region. For this purpose, we developed a method of analysis where complete, phase-solved, posterior-weighted haplotype assignments are imputed for each study individual from incomplete, multi-locus, genotyping data. The imputed assignments provide an input to a number of proposed procedures for testing association at a microsatellite level or of a sequence of SNPs. These include a Mantel-Haenszel type test based on expected frequencies of pseudocase/pseudocontrol haplotypes, as well as permutation based tests, including a combination of permutation and weighted logistic regression analysis. Application of these methods allowed us to find a significant association between MS and the SNP rs28936 located in the 3' UTR segment of ACCN1 with p = 0.0004 (p = 0.002, after adjusting for multiple testing). This result is in tune with several recent experimental findings which suggest that ACCN1 may play an important role in the pathogenesis of MS.     

Porin and cytochrome oxidase containing contact sites involved in the oxidation of cytosolic NADH

Cytochrome c (cyto-c) added to isolated mitochondria promotes the oxidation of extra-mitochondrial NADH and the reduction of molecular oxygen associated to the generation of an electrochemical membrane potential available for ATP synthesis. The electron transport pathway activated by exogenous cyto-c molecules is completely distinct from the one catalyzed by the respiratory chain. Dextran sulfate (500 kDa), known to interact with porin (the voltage-dependent anion channel), other than to inhibit the release of ATP synthesized inside the mitochondria, greatly decreases the activity of exogenous NADH/cyto-c system of intact mitochondria but has no effect on the reconstituted system made of mitoplasts and external membrane preparations. The results obtained are consistent with the existence of specific contact sites containing cytochrome oxidase and porin, as components of the inner and the outer membrane respectively, involved in the oxidation of cytosolic NADH. The proposal is put forward that the bi-trans-membrane electron transport chain activated by cytosolic cyto-c becomes, in physio-pathological conditions: (i) functional in removing the excess of cytosolic NADH; (ii) essential for cell survival in the presence of an impairment of the first three respiratory complexes; and (iii) an additional source of energy at the beginning of apoptosis.     

Effects of growth-promoting rhizobacteria on maize growth and rhizosphere microbial community under conservation tillage in Northeast China

Conservation tillage in conjunction with straw mulching is a sustainable agricultural approach. However, straw mulching reduces the soil temperature, inhibits early maize growth and reduces grain yield in cold regions. To address this problem, we investigated the effects of inoculation of plant growth-promoting rhizobacteria (PGPR) on maize growth and rhizosphere microbial communities under conservation tillage in Northeast China. The PGPR strains Sinorhizobium sp. A15, Bacillus sp. A28, Sphingomonas sp. A55 and Enterobacter sp. P24 were isolated from the maize rhizosphere in the same area and inoculated separately. Inoculation of these strains significantly enhanced maize growth, and the strains A15, A28 and A55 significantly increased grain yield by as much as 22%–29%. Real-time quantitative PCR and high-throughput sequencing showed that separate inoculation with the four strains increased the abundance and species richness of bacteria in the maize rhizosphere. Notably, the relative abundance of Acidobacteria_Subgroup_6, Chloroflexi_KD4-96, and Verrucomicrobiae at the class level and Mucilaginibacter at the genus level were positively correlated with maize biomass and yield. Inoculation with PGPR shows potential for improvement of maize production under conservation tillage in cold regions by regulating the rhizosphere bacterial community structure and by direct stimulation of plant growth.     

Maize based diets and possible neurobehavioural after-effects among some populations in the world

Maize is a cereal particularly lacking in tryptophan, which is the precursor of serotonin, an important neurotransmitter. Altough complementary foods may eliminate tryptophan deficiency, serotonin deficiency may often continue to exist because of competition made by other Large Neutral Amino Acids (LNAA) against tryptophan for neuron access, since they use the same carrier to cross the blood-brain barrier. Thus serotonin synthesis depends on two variables: the amount of tryptophan and the trp/LNAA ratio (R). “R” is lowest for common maize, low for beans and, as a rule, for most vegetable foods, higher for meat. So, when maize is the preponderant food in the meal, the “R” value lowers and so in parallel serotonin synthesis does. Serotonin deficiency involves several behavioural consequences, such as the tendency towards aggressive behaviour or the religious fanaticism. Among native american populations, these consequences appear, as a rule, positively correlated with maize alimentary dependence (Aztecs appear as those who greatly suffered from serotonin deficiency). In the world these are thinkable for some african populations (i.e. Zulu) or european (i.e. Balkan peoples).     

Influence of the denticity of ligand systems on the in vitro and in vivo behavior of (99m)Tc(I)-tricarbonyl complexes: a hint for the future functionalization of biomolecules

Functionalization of biologically relevant molecules for the labeling with the novel fac-[(99m)Tc(OH(2))(3)(CO)(3)](+) precursor has gained considerable attention recently. Therefore, we tested seven different tridentate (histidine L(1)(), iminodiacetic acid L(2)(), N-2-picolylamineacetic acid L(3)(), N, N-2-picolylaminediacetic acid L(4)()) and bidentate (histamine L(5)(), 2-picolinic acid L(6)(), 2,4-dipicolinic acid L(7)()) ligand systems, with the potential to be bifunctionalized and attached to a biomolecule. The ligands allowed mild radiolabeling conditions with fac-[(99m)Tc(OH(2))(3)(CO)(3)](+) (30 min, 75 degrees C). The ligand concentrations necessary to obtain yields of >95% of the corresponding organometallic complexes 1-7 ranged from 10(-)(6) to 10(-)(4) M. Complexes of the general formula "fac-[(99m)TcL(CO)(3)]" (L = tridentate ligand) and "fac-[(99m)Tc(OH(2))L'(CO)(3)]" (L' = bidentate ligand), respectively, were produced. Challenge studies with cysteine and histidine revealed significant displacement of the ligands in complexes 5-7 but only little exchange with complexes 1-4 after 24 h at 37 degrees C in PBS buffer. However, no decomposition to (99m)TcO(4)(-) was observed under these conditions. All complexes showed a hydrophilic character (log P(o/w) values ranging from -2.12 to 0.32). Time-dependent FPLC analyses of compounds 1-7 incubated in human plasma at 37 degrees C showed again no decomposition to (99m)TcO(4)(-) after 24 h at 37 degrees C. However, the complexes with bidentate ligands (5-7) became almost completely protein bound after 60 min, whereas the complexes with tridentate coordinated ligands (1-4) showed no reaction with serum proteins. The compounds were tested for their in vivo stability and the biodistribution characteristics in BALB/c mice. The complexes with tridentate coordinated ligand systems (1-4) revealed generally a good and fast clearance from all organs and tissues. On the other hand, the complexes with only bidentate coordinated ligands (5-7) showed a significantly higher retention of activity in the liver, the kidneys, and the blood pool. Detailed radiometric analyses of murine plasma samples, 30 min p.i. of complex fac-[(99m)TcL(1)(CO)(3)], 1, revealed almost no reaction of the radioactive complex with the plasma proteins. By contrast, in plasma samples of mice, which were injected with complex fac-[(99m)Tc(OH(2))L(5)(CO)(3)](+), 5, the entire radioactivity coeluded with the proteins. On the basis of these in vitro and in vivo experiments, it appears that functionalization of biomolecules with tridentate-chelating ligand systems is preferable for the labeling with fac-[(99m)Tc(OH(2))(3)(CO)(3)](+), since this will presumably result in radioactive bioconjugates with better pharmacokinetic profiles.     

Immunological functions of leptin and adiponectin

Recent years have seen several advances in our understanding of the functions of adipose tissue regarding not only the energy storage, but also the regulation of complex metabolic and endocrine functions. In this context, leptin and adiponectin, the two most abundant adipocyte products, represent one of the best example of adipocytokines involved in the control of energy expenditure, lipid and carbohydrate metabolism as well as in the regulation of immune responses. Leptin and adiponectin secretion is counter-regulated in vivo, in relation to degree of adiposity, since plasma leptin concentrations are significantly elevated in obese subjects in proportion to body mass index while adiponectin secretion decreases in relation to the amount of adipose tissue. In this review we focus on the main biological activities of leptin and adiponectin on the lipid and carbohydrate metabolism and on their contribute in regulation of innate and adaptive immune responses.     

Large and dissimilar repertoire of Melan-A/MART-1-specific CTL in metastatic lesions and blood of a melanoma patient

It is widely accepted that the repertoire of Melan-A-specific T cells naturally selected in melanoma patients is diverse and mostly nonoverlapping among different individuals. To date, however, no studies have addressed the TCR profile in different tumor sites and the peripheral blood from the same patient. We compared the TCR usage of Melan-A-specific T cells from different compartments of a single melanoma patient to evaluate possible clonotype expansion or preferential homing over a 4-mo follow-up period. Using HLA-A2 peptide tetramers, CD8(+) T cells recognizing the modified Melan-A immunodominant ELAGIGILTV peptide were isolated from four metastatic lesions resected from a single melanoma patient, and their TCR repertoire was studied. A panel of T cell clones was generated by cell cloning of tetramer-positive cells. Analysis of the TCR beta-chain V segment and the complementarity-determining region 3 (CDR3) length and sequence revealed a large diversity in the TCR repertoire, with only some of the clones showing a partial conservation in the CDR3. A similar degree of diversity was found by analyzing a number of T cell clones obtained after sorting a Melan-A-specific population derived from PBLs of the same patient after in vitro culture with the immunodominant epitope. Moreover, clonotypes found at one site were not present in another, suggesting the lack of expansion and circulation of one or more clonotypes. Taken together, these results buttress the notion that the CTLs recognizing the immunodominant Ag of Melan-A comprise a high number of different clonotypic TCR, of which only some exhibit common features in the CDR3.