The Effect of Age and Gender on 38 Chemical Element Contents in Human Femoral Neck Investigated by Instrumental Neutron Activation Analysis

In the present study, the association of arsenic, cadmium, lead, and nickel between scalp hair samples and mortality from myocardial infarction (MI) patients at first, second, and third heart attack was studied. The biological samples of 130 MI patients (77 male and 53 female) age ranged (45–60 years), were collected and 61 healthy persons of same age group (33 male and 28 female) was selected as control subjects. The toxic elements (TEs) in biological samples were assessed by the electrothermal atomic absorption spectrophotometry prior to microwave-assisted acid digestion. The validity of methodology was checked by the certified human hair reference material (BCR 397). It was observed during the study that 78% of 32 patients of the third MI, age >50 years were expired. In these subjects, the level of As, Cd, Ni, and Pb were increased by 10.6%, 19.5%, 15.7%, and 9.8% in the scalp hair as compared to those who tolerated third MI attack (p = 0.12). The high level of toxic metals may play a role in the development of heart disease in the subjects of this study.     

Effects of aging and gender on the spatial organization of nuclei in single human skeletal muscle cells

The skeletal muscle fibre is a syncitium where each myonucleus regulates the gene products in a finite volume of the cytoplasm, i.e., the myonuclear domain (MND). We analysed aging‐ and gender‐related effects on myonuclei organization and the MND size in single muscle fibres from six young (21–31 years) and nine old men (72–96 years), and from six young (24–32 years) and nine old women (65–96 years), using a novel image analysis algorithm applied to confocal images. Muscle fibres were classified according to myosin heavy chain (MyHC) isoform expression. Our image analysis algorithm was effective in determining the spatial organization of myonuclei and the distribution of individual MNDs along the single fibre segments. Significant linear relations were observed between MND size and fibre size, irrespective age, gender and MyHC isoform expression. The spatial organization of individual myonuclei, calculated as the distribution of nearest neighbour distances in 3D, and MND size were affected in old age, but changes were dependent on MyHC isoform expression. In type I muscle fibres, average NN‐values were lower and showed an increased variability in old age, reflecting an aggregation of myonuclei in old age. Average MND size did not change in old age, but there was an increased MND size variability. In type IIa fibres, average NN‐values and MND sizes were lower in old age, reflecting the smaller size of these muscle fibres in old age. It is suggested that these changes have a significant impact on protein synthesis and degradation during the aging process.     

Dysgonomonas hofstadii sp. nov., isolated from a human clinical source

A Gram-negative staining, facultative anaerobic, cocco-bacillus-shaped organism was isolated from a post-operative abdominal wound. Based on morphological and biochemical criteria, strain MX 1040 ( = CCUG 54731^T) was tentatively identified as Bacteroidaceae but did not correspond to any recognized species of this family. Comparative 16S rRNA gene sequencing analysis demonstrated the organism to be related to species of the genus Dysgonomonas, although sequence divergence values of >5% with the other members of this genus demonstrated the organism to represent a novel species. Phylogenetic analysis revealed the novel organism to be most closely related to Dysgonomonas gadei. The major long-chain cellular fatty acids of the novel species consisted of iso-C_14:0, anteiso-C_15:0, C_16:0, and iso-C_16:0. Based on the phenotypic criteria and phylogenetic considerations, it is proposed that strain MX 1040 from a human clinical source represents a new species of the genus Dysgonomonas, as Dysgonomonas hofstadii sp. nov. The type strain of D. hofstadii is CCUG 54731^T ( = CCM 7606^T).     

Characterization of the Desulfovibrio desulfuricans ATCC 27774 DsrMKJOP complex--a membrane-bound redox complex involved in the sulfate respiratory pathway

Sulfate-reducing organisms use sulfate as an electron acceptor in an anaerobic respiratory process. Despite their ubiquitous occurrence, sulfate respiration is still poorly characterized. Genome analysis of sulfate-reducing organisms sequenced to date permitted the identification of only two strictly conserved membrane complexes. We report here the purification and characterization of one of these complexes, DsrMKJOP, from Desulfovibrio desulfuricans ATCC 27774. The complex has hemes of the c and b types and several iron-sulfur centers. The corresponding genes in the genome of Desulfovibrio vulgaris were analyzed. dsrM encodes an integral membrane cytochrome b; dsrK encodes a protein homologous to the HdrD subunit of heterodisulfide reductase; dsrJ encodes a triheme periplasmic cytochrome c; dsrO encodes a periplasmic FeS protein; and dsrM encodes another integral membrane protein. Sequence analysis and EPR studies indicate that DsrJ belongs to a novel family of multiheme cytochromes c and that its three hemes have different types of coordination, one bis-His, one His/Met, and the third a very unusual His/Cys coordination. The His/Cys-coordinated heme is only partially reduced by dithionite. About 40% of the hemes are reduced by menadiol, but no reduction is observed upon treatment with H2 and hydrogenase, irrespective of the presence of cytochrome c3. The aerobically isolated Dsr complex displays an EPR signal with similar characteristics to the catalytic [4Fe-4S]3+ species observed in heterodisulfide reductases. Further five different [4Fe-4S](2+/1+) centers are observed during a redox titration followed by EPR. The role of the DsrMKJOP complex in the sulfate respiratory chain of Desulfovibrio spp. is discussed.     

Screening Possible Mechanisms Mediating Cadmium Resistance in Rhizobium leguminosarum bv. viciae Isolated from Contaminated Portuguese Soils

Environment heavy-metal contamination is now widespread. Soils may become contaminated from a variety of anthropogenic sources, such as smelters, mining, industry, and application of metal-containing pesticides and fertilizers. Soil microorganisms are very sensitive to moderate heavy-metal concentrations. Therefore, the present work was designed to screen possible mechanisms involved in Rhizobium's Cd resistance; with this purpose, we determined the tolerance levels of several isolates originated from sites with different heavy-metal contamination. Whole-cell-soluble proteins and plasmid profiles were analyzed. We also determined Cd cell concentrations and lipopolysaccharide (LPS) amounts. Results showed different tolerances among Rhizobium isolates; according to their maximum resistance level, isolates were divided in four groups: sensitive (0–125 μM CdCl₂), moderately tolerant (125–210 μM CdCl₂), tolerant (250–500 μM CdCl₂), and extremely tolerant (≥750 μM CdCl₂). Intracellular Cd concentrations were lower when compared to wall-bound Cd. Unexpectedly, extremely tolerant isolates accumulated higher levels of metal, suggesting the presence of intracellular agents that prevent metal interfering with important metabolic pathways. The electrophoretic patterns of whole-cell-soluble proteins evidenced cadmium as an inducer of protein metabolism alterations, which were more evident in some polypeptides. Plasmid profiles also showed differences; most tolerant isolates presented two plasmids with molecular weights of 485 and 415 kb, indicating that extrachromosomal DNA may be involved in cadmium resistance. LPS showed to be a common mechanism of resistance. However, the degree of tolerance conferred by LPS is not enough to support tolerance to the higher levels of stress imposed. Presence of other resistance mechanisms is currently being investigated.     

Isolation and characterisation of a cDNA encoding a novel cytosolic ascorbate peroxidase from potato plants (Solanum tuberosum L.)

Ascorbate peroxidase (APX) is one of the key enzymes of the plant antioxidant system playing, along with catalase, a central role in hydrogen peroxide scavenging. An approach to further increase the knowledge about cytosolic APX gene organization can be achieved by isolating and characterisating new cDNAs, thus providing new insights about the physiological roles and regulation of these enzymes. A partial cDNA clone (corresponding to the 3’ untranslated region), cytosolic ascorbate peroxidase-related, was isolated from potato sprouts by RT-PCR. Database analysis retrieved several expressed sequence tags (ESTs) coding potato cytosolic ascorbate peroxidase, that were used to infer the complete cDNA sequence. The deduced amino acid sequence revealed high homologies with other plant cytosolic ascorbate peroxidases, confirming the reliability of the virtual cDNA. Northern blot analysis revealed the existence of a single band related to the isolated cDNA and the southern blotting results allowed the elaboration of a possible gene organization.     

Evaluation of the presence of aspartic proteases from Centaurea calcitrapa during seed germination

Aspartic proteinases are present in a variety of organisms including plants. Common features of aspartic proteases include an active site cleft that contains two catalytic aspartic residues, acid pH optima for enzymatic activity, inhibition by pepstatin A. Plant aspartic proteinases occur in seeds and may be involved in the processing of storage proteins. Many of them have been purified and characterized. The presence of aspartic proteases in seeds of Centaurea calcitrapa during germination was investigated by measuring the activity on enzyme extracts. The aspartic proteases are present mainly in the beginning of seed germination suggesting that they could initiate the degradation of protein reserves in germinating seeds.

These proteases were purified by salt precipitation followed by anion-exchange chromatography. Purified aspartic proteases have an optimal pH between 3.5 and 4.5, using FTC-hemoglobin as substrate and an optimal temperature at 52 °C. The ability of seed extracts for milk clotting was tested and the clotting time that was achieved is in the same range found for flower extracts appropriated for special cheeses in which weak clotting agents are required.     

Identification of new central nervous system specific mouse microRNAs

MicroRNAs (miRNAs) are small regulatory molecules suppressing mRNA activity in metazoans. Here we describe two new miRNAs cloned from brain tissue of mouse embryos. These miRNAs are expressed mainly during embryogenesis and specifically in the central nervous system. We also established the expression patterns of three recently identified miRNAs that were found in our short RNA library. All of them were expressed in the brain and spinal chord but while miR-410 and miR-431 were central nervous system specific, miR-500 was also expressed in limb buds. In addition, the expression of miR-500 in limb buds showed very strong asymmetry in favour of the left hand side.